Protease inhibitors partially overcome the interferon nonresponse phenotype in patients with chronic hepatitis C.

The outcome of triple therapy with protease inhibitors (PI) depends on the intrinsic response to interferon. Interferon-stimulated gene (ISG) expression differs by cell type in the liver and is a strong predictor of interferon responsiveness. Patients who respond well to interferon have low/absent ISG expression in hepatocytes but significant ISG expression in macrophages. Nonresponders (NRs) show the opposite pattern. We aimed to determine the association between cell-type-specific ISG staining and treatment outcome with PI-based triple therapy. Liver biopsy tissue from consecutive patients treated with boceprevir or telaprevir with peginterferon and ribavirin was stained for myxovirus A (MxA). Staining was scored 0-3 in macrophages (M-MxA) and hepatocytes (H-MxA), and IL28B genotyping was performed. Of 56 patients included 41 achieved SVR (73%) (sustained virological response), 2 (4%) relapsed, 10 (18%) were NRs, and 3 (5%) were lost to follow-up. Median M-MxA staining was stronger and H-MxA staining was weaker in patients who achieved SVR. MxA staining correlated with IL28B genotype and with the HCV RNA decline during lead-in phase. However, unlike with dual therapy, the negative predictive value (NPV) of absent or weak M-MxA staining was poor (42%), while the positive predictive value improved (93%). Although by multivariable logistic regression M-MxA staining was significantly associated with SVR (OR 4.35, 1.32-14.28, P = 0.012), the predictive ability was inadequate to withhold therapy. The interaction between macrophages and hepatocytes plays a critical role in interferon responsiveness; however, the addition of a PI at least partially overcomes the interferon nonresponse phenotype making the predictive ability of ISG staining less clinically useful.

Three-dimensional characterization of resorption cavity size and location in human vertebral trabecular bone.

The number and size of resorption cavities in cancellous bone are believed to influence rates of bone loss, local tissue stress and strain and potentially whole bone strength. Traditional two-dimensional approaches to measuring resorption cavities in cancellous bone report the percent of the bone surface covered by cavities or osteoclasts, but cannot measure cavity number or size. Here we use three-dimensional imaging (voxel size 0.7×0.7×5.0 µm) to characterize resorption cavity location, number and size in human vertebral cancellous bone from nine elderly donors (7 male, 2 female, ages 47-80 years). Cavities were 30.10 ± 8.56 µm in maximum depth, 80.60 ± 22.23∗10(3) µm(2) in surface area and 614.16 ± 311.93∗10(3) µm(3) in volume (mean ± SD). The average number of cavities per unit tissue volume (N.Cv/TV) was 1.25 ± 0.77 mm(-3). The ratio of maximum cavity depth to local trabecular thickness was 30.46 ± 7.03% and maximum cavity depth was greater on thicker trabeculae (p<0.05, r(2)=0.14). Half of the resorption cavities were located entirely on nodes (the intersection of two or more trabeculae) within the trabecular structure. Cavities that were not entirely on nodes were predominately on plate-like trabeculae oriented in the cranial-caudal (longitudinal) direction. Cavities on plate-like trabeculae were larger in maximum cavity depth, cavity surface area and cavity volume than cavities on rod-like trabeculae (p<0.05). We conclude from these findings that cavity size and location are related to local trabecular microarchitecture.

Mechanical failure begins preferentially near resorption cavities in human vertebral cancellous bone under compression.

The amount of bone turnover in the body has been implicated as a factor that can influence fracture risk and bone strength. Here we test the idea that remodeling cavities promote local tissue failure by determining if microscopic tissue damage (microdamage) caused by controlled loading in vitro is more likely to form near resorption cavities. Specimens of human vertebral cancellous bone (L4, 7 male and 2 female, age 70±10, mean±SD) were loaded in compression to the yield point, stained for microscopic tissue damage and submitted to three-dimensional fluorescent imaging using serial milling (image voxel size 0.7×0.7×5.0 µm). We found the resulting damage volume per bone volume (DV/BV) was correlated with percent eroded surface (p<0.01, r(2)=0.65), demonstrating that whole specimen measures of resorption cavities and microdamage are related. Locations of microdamage were more than two times as likely to have a neighboring resorption cavity than randomly selected sites without microdamage (relative risk 2.39, 95% confidence interval of relative risk: 2.09-2.73), indicating a spatial association between resorption cavities and microdamage at the local level. Individual microdamage sites were 48,700 (40,100; 62,700) µm(3) in size (median, 25th and 75th percentiles). That microdamage was associated with resorption cavities when measured at the whole specimen level as well as at the local level provides strong evidence that resorption cavities play a role in mechanical failure processes of cancellous bone and therefore have the potential to influence resistance to clinical fracture.

Liver allograft pathology: approach to interpretation of needle biopsies with clinicopathological correlation.

The spectrum of diseases encountered in post-transplant liver pathology biopsies is broad. In this review, these have been divided as belonging to one of three categories: (1) new-onset/de novo post-transplant abnormalities (early and late), (2) rejection, and (3) recurrence of original disease. The clinical and pathological features of the entities making up each category, with the relevant differential diagnosis and overlaps between and within these groups, are discussed and illustrated. Recurrent or de novo neoplasms make up a fourth category not included in this review. Early new-onset conditions are mostly related to surgical complications, donor factors and ischaemia to the graft. These include reperfusion/preservation injury, lipopeliosis, small-for-size-syndrome, biliary sludge syndrome and hepatic artery thrombosis. The various forms of rejection (cellular, chronic, antibody-mediated, and late atypical rejection) are detailed. Most chronic liver diseases can and do recur in the graft. They may display features that overlap with de novo conditions (eg, primary sclerosing cholangitis versus chronic rejection). As with most cases of allograft biopsy interpretation, accurate diagnosis rests with careful correlation of histological features with clinical, imaging and laboratory findings, and often comparison with previous sequential and follow-up biopsies. Late-onset new diseases include biliary strictures, idiopathic chronic hepatitis and de novo autoimmune hepatitis, among others. This review provides a practical approach to the interpretation of these challenging biopsies. Selected difficult scenarios or conundrums are identified and discussed in the relevant sections.

Gastric mucin is expressed in a subset of endocervical tunnel clusters: type A tunnel clusters of gastric phenotype.

AIMS: Gastric mucin expression has been demonstrated in a group of endocervical glandular lesions. The aim of this study was to gain further insight into endocervical lesions with a gastric phenotype. METHODS AND RESULTS: Various types of tunnel clusters (TC) were examined for gastric mucin by alcian blue/periodic acid-Schiff staining and immunohistochemistry for HIK1083. Five of 34 cases of TC expressed gastric mucin defined by PAS dominant neutral mucin and immunopositivity for pyloric gland mucin. Histologically, TC expressing gastric mucin showed lobular arrangements of small to medium-sized glands composed of mucin-rich columnar cells and were classified as Flumann's type A TC. Neither type B TC nor normal endocervical glands expressed PAS dominant neutral mucin and none of them was immunopositive for pyloric gland mucin. Five patients with type A TC of gastric phenotype ranged in age from 33 to 79 years (mean 58 years) and were multiparous. Type A TC of gastric phenotype, ranging from 2 to 4 mm in maximum diameter, were incidental findings in hysterectomy specimens. CONCLUSION: Type A TC of gastric phenotype could be related to lobular endocervical glandular hyperplasia of gastric phenotype. The pathogenesis of gastric metaplasia in TC remains unclear.

Dicer functions in RNA interference and in synthesis of small RNA involved in developmental timing in C. elegans.

Double-stranded RNAs can suppress expression of homologous genes through an evolutionarily conserved process named RNA interference (RNAi) or post-transcriptional gene silencing (PTGS). One mechanism underlying silencing is degradation of target mRNAs by an RNP complex, which contains approximately 22 nt of siRNAs as guides to substrate selection. A bidentate nuclease called Dicer has been implicated as the protein responsible for siRNA production. Here we characterize the Caenorhabditis elegans ortholog of Dicer (K12H4.8; dcr-1) in vivo and in vitro. dcr-1 mutants show a defect in RNAi. Furthermore, a combination of phenotypic abnormalities and RNA analysis suggests a role for dcr-1 in a regulatory pathway comprised of small temporal RNA (let-7) and its target (e.g., lin-41).

Regulated transposition of a fish transposon in the mouse germ line.

Tc1/mariner elements are able to transpose in species other than the host from which they were isolated. As potential vectors for insertional mutagenesis and transgenesis of the mouse, these cut-and-paste transposons were tested for their ability to transpose in the mouse germ line. First, the levels of activity of several Tc1/mariner elements in mammalian cells were compared; the reconstructed fish transposon Sleeping Beauty (SB) was found to be an order of magnitude more efficient than the other tested transposons. SB then was introduced into the mouse germ line as a two-component system: one transgene for the expression of the transposase in the male germ line and a second transgene carrying a modified transposon. In 20% of the progeny of double transgenic male mice the transposon had jumped from the original chromosomal position into another locus. Analysis of the integration sites shows that these jumps indeed occurred through the action of SB transposase, and that SB has a strong preference for intrachromosomal transposition. Analysis of the excision sites suggests that double-strand breaks in haploid spermatids are repaired via nonhomologous end joining. The SB system may be a powerful tool for transposon mutagenesis of the mouse germ line.

Measurement of the gadopentetate dimeglumine partition coefficient in human myocardium in vivo: normal distribution and elevation in acute and chronic infarction.

PURPOSE: To establish a method for measuring the partition coefficient (lambda) of gadopentetate dimeglumine in humans in vivo, evaluate the spatial and intersubject variation in the lambda of normal myocardium, and compare these values on a regional basis with lambda values of acute and chronic infarcted myocardium. MATERIALS AND METHODS: Twelve healthy subjects and patients with acute (n = 5) or chronic (n = 5) myocardial infarction underwent magnetic resonance imaging at 1.5 T. Look-Locker images were acquired at four short-axis levels to measure myocardial and blood longitudinal relaxation time at baseline and after a 30-40-minute infusion of gadopentetate dimeglumine. lambda was calculated as DeltaR1(M)/DeltaR1(B, )where M = myocardium, and B = blood. RESULTS: The magnitude of the estimated lambda in normal myocardium was uniform over the entire myocardium at 0.56 mL/g +/- 0.10 (SD). The lambda values in patients with acute (0.91 mL/g +/- 0.11, P <.001) or chronic (lambda = 0.78 mL/g +/- 0.09, P <.001) infarction were significantly elevated, as compared with those in healthy subjects. A 20% elevation in lambda, as compared with the mean value of a corresponding normal circumferential segment, allowed identification of chronically (sensitivity, 88%; specificity, 96%) or acutely (sensitivity, 100%; specificity, 98%) infarcted segments. CONCLUSION: Quantification of the lambda in vivo allows differentiation between normal and acutely or chronically infarcted myocardium, with high sensitivity and specificity.

Performance of QRS detection for cardiac magnetic resonance imaging with a novel vectorcardiographic triggering method.

In many cardiac patients, image quality and/or scan efficiency is reduced due to imprecise R-wave ability to trigger the scan due to noise on the electrocardiogram (ECG) caused by the magnetic resonance (MR) environment. We developed a triggering system that uses the spatial information of the vectorcardiogram (VCG) to minimize the effects of MR-related noise on triggering. Fifteen volunteers underwent standard cardiovascular MR exams, and a total of 52,474 R-waves were evaluated with the algorithm, giving a performance index of 99.91%. The mean propagation delay of the system was -10.64 +/- 3.19 msec, which falls within the real-time definition for cardiac MRI triggering. Five patients had arrhythmias consisting of premature ventricular depolarizations (PVDs) and supraventricular extra systoles. For those patients with PVDs, all arrhythmic beats were rejected unless they passed through the algorithm's reference point. The performance index for the arrhythmic patients approached 100%. VCG-based triggering has been demonstrated to provide near 100% triggering performance during cardiac MR examinations.

Noninvasive modalities. Cardiac MR imaging.

As outlined in this article, the strength of MR imaging is that it can provide flow, function, and in some cases metabolic data in a single examination, independent of patient body habitus. Future prospects for real-time imaging and in vivo mapping of fiber orientation promise further advances in our understanding of the structure-function relationship in diastole. Many of the MR imaging methods that have been developed for cardiovascular imaging are now mature and available on state-of-the-art scanners. Although MR imaging can provide detailed characterization of diastolic function, there is a paucity of clinical results which could lead to use guidelines. When more clinicians have access and become familiar with MR imaging, and the type of information that it can provide, clinical trials will be needed to establish the role of MR imaging for evaluation of diastolic function. In the meantime, MR imaging remains an excellent research tool for this application and will help yield further insights into the pathophysiology of diastolic dysfunction.

Quantification of left ventricular function with magnetic resonance images acquired in real time.

The application of real-time magnetic resonance imaging (MRI) techniques to cardiac imaging is particularly attractive because current MR examinations of left ventricular (LV) function can be prohibitively long and are dependent on electrocardiographic triggering. We conducted a study of the minimum spatial and temporal resolution requirements necessary for real-time ventricular function MR imaging to quantify LV volumes accurately, both at resting conditions and during cardiac stress tests. In addition, we implemented a real-time segmented echoplanar imaging pulse sequence and used it to quantify LV volume in 10 healthy volunteers. We compared these results with those obtained using conventional gradient-echo cine imaging and found good agreement throughout the cardiac cycle (mean difference -0.8 +/- 10.6 ml). In conclusion, real-time cardiac MR imaging can be used to quantify LV volumes accurately throughout the cardiac cycle, over the physiologic range of heart rates, thereby decreasing the time required for a complete functional cardiac examination. J. Magn. Reson. Imaging 2000;12:430-438.

Left ventricular contractility is impaired following myocardial infarction in the pig and rat: assessment by the end systolic pressure-volume relation using a single-beat estimation technique and cine magnetic resonance imaging.

The end systolic pressure-volume relation (ESPVR) has been shown to be a relatively load independent measure of left ventricular (LV) contractility. Recently, several single-beat ESPVR computation methods have been developed, enabling the quantification of LV contractility without the need to alter vascular loading conditions on the heart. Using a single-beat ESPVR method, which has been validated previously in humans and assumes that normalized elastance is constant between individuals of a species, we studied the effects of myocardial infarction on LV contractility in two species, the rat and the pig. In our studies, LV pressure was acquired invasively and LV volume determined noninvasively with magnetic resonance imaging, at one week postinfarction in pigs and at 12 weeks postinfarction in rats. Normalized systolic elastance curves in both animal species were not statistically different from that of humans. Also, the slope of the ESPVR (Ees) decreased significantly following infarction in both species, while the volume-axis intercept (V0) was unaffected. These results indicate that a single-beat ESPVR method can be used to measure the inotropic response of the heart to myocardial infarction, and that the basis for this method (i.e., constant normalized elastance) is applicable to a variety of mammalian species.

Cardiac rotation and relaxation in patients with aortic valve stenosis.

BACKGROUND: Diastolic dysfunction with delayed relaxation and abnormal passive elastic properties has been described in patients with severe pressure overload hypertrophy. The purpose of this study was to evaluate the time course of rotational motion of the left ventricle in patients with aortic valve stenosis using myocardial tagging. METHODS: Myocardial tagging is a non-invasive method based on magnetic resonance which makes it possible to label ('tag') specific myocardial regions. From the motion of the tag's cardiac rotation, radial displacement and translational motion can be determined. In 12 controls and 13 patients with severe aortic valve stenosis systolic and diastolic wall motion was assessed in an apical and basal short axis plane. RESULTS: The normal left ventricle performs a systolic wringing motion around the ventricular long axis with clockwise rotation at the base (-4.4+/-1.6 degrees) and counter-clockwise rotation at the apex (+6.8+/-2.5 degrees) when viewed from the apex. During early diastole an untwisting motion can be observed which precedes diastolic filling. In patients with aortic valve stenosis systolic rotation is reduced at the base (-2.4+/-2.0 degrees; P<0.01) but increased at the apex (+12.0+/-6.0 degrees; P<0.05). Diastolic untwisting is delayed and prolonged with a decrease in normalized rotation velocity (-6.9+/-1.1 s(-1)) when compared to controls (-10.7+/-2.2 s(-1); P<0.001). Maximal systolic torsion is 8.0+/-2.1 degrees in controls and 14.1+/-6.4 degrees (P<0.01) in patients with aortic valve stenosis. CONCLUSIONS: Left ventricular pressure overload hypertrophy is associated with a reduction in basal and an increase in apical rotation resulting in increased torsion of the ventricle. Diastolic untwisting is delayed and prolonged. This may explain the occurrence of diastolic dysfunction in patients with severe pressure overload hypertrophy.

Single breath-hold slice-following CSPAMM myocardial tagging.

Myocardial tagging has shown to be a useful magnetic resonance modality for the assessment and quantification of local myocardial function. Many myocardial tagging techniques suffer from a rapid fading of the tags, restricting their application mainly to systolic phases of the cardiac cycle. However, left ventricular diastolic dysfunction has been increasingly appreciated as a major cause of heart failure. Subtraction based slice-following CSPAMM myocardial tagging has shown to overcome limitations such as fading of the tags. Remaining impediments to this technique, however, are extensive scanning times (approximately 10 min), the requirement of repeated breath-holds using a coached breathing pattern, and the enhanced sensitivity to artifacts related to poor patient compliance or inconsistent depths of end-expiratory breath-holds. We therefore propose a combination of slice-following CSPAMM myocardial tagging with a segmented EPI imaging sequence. Together with an optimized RF excitation scheme, this enables to acquire as many as 20 systolic and diastolic grid-tagged images per cardiac cycle with a high tagging contrast during a short period of sustained respiration.

Cis requirements for transposition of Tc1-like transposons in C. elegans.

The Caenorhabditis elegans transposons Tc1 and Tc3 are able to transpose in heterologous systems such as human cell lines and zebrafish. Because these transposons might be useful vectors for transgenesis and mutagenesis of diverse species, we determined the minimal cis requirements for transposition. Deletion mapping of the transposon ends shows that fewer than 100 bp are sufficient for transposition of Tc3. Unlike Tc1, Tc3 has a second, internal transposase binding site at each transposon end. We found that these binding sites play no major role in the transposition reaction, since they can be deleted without reduction of the transposition frequency. Site-directed mutagenesis was performed on the conserved terminal base pairs at the Tc3 ends. The four terminal base pairs at the ends of the Tc3 inverted repeats were shown to be required for efficient transposition. Finally, increasing the length of the transposon from 1.9 kb to 12.5 kb reduced the transposition frequency by 20-fold, both in vivo and in vitro.

Novel real-time R-wave detection algorithm based on the vectorcardiogram for accurate gated magnetic resonance acquisitions.

Electrocardiograph (ECG) triggered or gated magnetic resonance methods are used in many imaging applications. Therefore, a reliable trigger signal derived from to the R-wave of the ECG is essential, especially in cardiac imaging. However, currently available methods often fail mainly due to the artifacts in the ECG generated by the MR scanner itself, such as the magnetohydrodynamic effect and gradient switching noise. The purpose this study was to characterize the accuracy of selected R-wave detection algorithms in an MR environment, and to develop novel approaches to eliminate imprecise triggering. Vectorcardiograms (VCG) in 12 healthy volunteers exposed to 1.5 T magnetic field were digitized and used as a reference data set including manually corrected onsets of R-waves. To define the magnetohydrodynamic effect, the VCGs were characterized in time, frequency, and spatial domains. The selected real-time R-wave detection algorithms, and a new "target-distance" VCG-based algorithm were applied either to standard surface leads calculated from the recorded VCG or to the VCG directly. The flow related artifact was higher in amplitude than the R-wave in 28% of the investigated VCGs which yielded up to 9-16%false positive detected QRS complexes for traditional algorithms. The "target-distance" R-wave detection algorithm yielded a score of 100% for detection with 0.2% false positives and was superior to all the other selected methods. Thus, the VCG of subjects exposed to a strong magnetic field can be use to separate the magnetohydrodynamic artifact and the actual R-wave, and markedly improves the trigger accuracy in gated magnetic resonance scans. Magn Reson Med 42:361-370, 1999.

Alterations in the local myocardial motion pattern in patients suffering from pressure overload due to aortic stenosis.

BACKGROUND: MR tissue tagging allows the noninvasive assessment of the locally and temporally resolved motion pattern of the left ventricle. Alterations in cardiac torsion and diastolic relaxation of the left ventricle were studied in patients with aortic stenosis and were compared with those of healthy control subjects and championship rowers with physiological volume-overload hypertrophy. METHODS AND RESULTS: Twelve aortic stenosis patients, 11 healthy control subjects with normal left ventricular function, and 11 world-championship rowers were investigated for systolic and diastolic heart wall motion on a basal and an apical level of the myocardium. Systolic torsion and untwisting during diastole were examined by use of a novel tagging technique (CSPAMM) that provides access to systolic and diastolic motion data. In the healthy heart, the left ventricle performs a systolic wringing motion, with a counterclockwise rotation at the apex and a clockwise rotation at the base. Apical untwisting precedes diastolic filling. In the athlete's heart, torsion and untwisting remain unchanged compared with those of the control subjects. In aortic stenosis patients, torsion is significantly increased and diastolic apical untwisting is prolonged compared with those of control subjects or athletes. CONCLUSIONS: Torsional behavior as observed in pressure- and volume-overloaded hearts is consistent with current theoretical findings. A delayed diastolic untwisting in the pressure-overloaded hearts of the patients may contribute to a tendency toward diastolic dysfunction.

Benefit of T1 reduction for magnetic resonance coronary angiography: a numerical simulation and phantom study.

Contrast agents have dramatically improved magnetic resonance angiography (MRA) of the abdominal and peripheral arteries. The imaging technique for these applications is usually a steady-state acquisition, for which the relationship between T1 in blood and the MR signal is well known. However, in electrocardiography-triggered angiography with limited acquisition windows, this relationship is more complex. Therefore the purpose of this work is to define the relationship between the T1 in blood and the MR signal amplitude in three-dimensional magnetic resonance coronary angiography (3D-MRCA). Simulations were performed using equations describing the MR signal in both steady-state and triggered acquisition schemes. Triggered acquisition schemes use flip-angle sweeps to maintain a constant signal during the acquisition. In this study, the effect of the flip angle sweep was calculated as a function of T1. The results show that the effect of T1 shortening in contrast-enhanced 3D-MRCA differs substantially from that in conventional contrast-enhanced MRA. The triggered acquisition allows unsaturated blood to enter the volume between the acquisitions and thereby gives a much higher signal at long T1s than does steady-state acquisition. Therefore, to gain a benefit in signal amplitude with contrast agents for 3D-MRCA using gradient-echo sequences, the T1 in blood may have to be as low as 50 msec. In addition, when using a prepulse to null myocardium, the results indicate the need for a large difference in T1 between blood and myocardium to avoid signal loss in blood.

Toward high-resolution myocardial tagging.

Magnetic resonance imaging with preceding tissue tagging is a robust method for assessing cardiac motion of the entire heartbeat cycle with a high degree of accuracy. One limitation of this technique, however, is the low resolution of the obtained displacement map of the labeled points within the myocardium. By a new tagging technique, which is based on the combination of two or more measurements of the same slice but with different grid positions, a highly improved resolution of cardiac motion data can be achieved. In combination with a multi-heart-phase echo-planar imaging sequence, such images with doubled grid frequency can be acquired in two short breath-hold periods.

Blood pool agent strongly improves 3D magnetic resonance coronary angiography using an inversion pre-pulse.

The ability of a blood pool contrast agent to enhance MR coronary angiography was defined. The proximal coronary vessels of pigs were imaged before and after administration of Gd-DTPA bound covalently to bovine serum albumin (0.2 mmol/ kg). The contrast agent resulted in a reduction of the blood T1 value to 33+/-5 msec, as determined in vivo with a Look-Locker technique. Both 2D and 3D imaging techniques were performed. An inversion pulse suppressed the signal of nonblood tissue postcontrast. After contrast agent administration, in the 3D data set the signal-to-noise ratio (SNR) of blood and contrast-to-noise ratio (CNR) of blood to myocardium were improved by factors of 2.0+/-0.2 and 15+/-8, respectively (P < 0.05). Postcontrast, the 3D acquisition was superior to the 2D technique in terms of spatial resolution, SNR of blood, and CNR of blood to myocardium. The high contrast of the 3D data set allowed for direct and rapid display of coronary arteries using a "closest vessel projection."