The ATPase activity of MLH1 is required to orchestrate DNA double-strand breaks and end processing during class switch recombination.

PublishedJournal ArticleResearch Support, N.I.H., ExtramuralResearch Support, Non-U.S. Gov'tAntibody diversification through somatic hypermutation (SHM) and class switch recombination (CSR) are similarly initiated in B cells with the generation of U:G mismatches by activation-induced cytidine deaminase but differ in their subsequent mutagenic consequences. Although SHM relies on the generation of nondeleterious point mutations, CSR depends on the production of DNA double-strand breaks (DSBs) and their adequate recombination through nonhomologous end joining (NHEJ). MLH1, an ATPase member of the mismatch repair (MMR) machinery, is emerging as a likely regulator of whether a U:G mismatch progresses toward mutation or DSB formation. We conducted experiments on cancer modeled ATPase-deficient MLH1G67R knockin mice to determine the function that the ATPase domain of MLH1 mediates in SHM and CSR. Mlh1(GR/GR) mice displayed a significant decrease in CSR, mainly attributed to a reduction in the generation of DSBs and diminished accumulation of 53BP1 at the immunoglobulin switch regions. However, SHM was normal in these mice, which distinguishes MLH1 from upstream members of the MMR pathway and suggests a very specific role of its ATPase-dependent functions during CSR. In addition, we show that the residual switching events still taking place in Mlh1(GR/GR) mice display unique features, suggesting a role for the ATPase activity of MLH1 beyond the activation of the endonuclease functions of its MMR partner PMS2. A preference for switch junctions with longer microhomologies in Mlh1(GR/GR) mice suggests that through its ATPase activity, MLH1 also has an impact in DNA end processing, favoring canonical NHEJ downstream of the DSB. Collectively, our study shows that the ATPase domain of MLH1 is important to transmit the CSR signaling cascade both upstream and downstream of the generation of DSBs.Spanish Ministry of Education and ScienceNIHNational Women’s Division of the Albert Einstein College of Medicin

Accelerated MRI at 9.4 T with electronically modulated time-varying receive sensitivities

PURPOSE To investigate how electronically modulated time-varying receive sensitivities can improve parallel imaging reconstruction at ultra-high field. METHODS Receive sensitivity modulation was achieved by introducing PIN diodes in the receive loops, which allow rapid switching of capacitances in both arms of each loop coil and by that alter B$_{1}$ $^{-}$ profiles, resulting in two distinct receive sensitivity configurations. A prototype 8-channel reconfigurable receive coil for human head imaging at 9.4T was built, and MR measurements were performed in both phantom and human subject. A modified SENSE reconstruction for time-varying sensitivities was formulated, and g-factor calculations were performed to investigate how modulation of receive sensitivity profiles during image encoding can improve parallel imaging reconstruction. The optimized modulation pattern was realized experimentally, and reconstructions with the time-varying sensitivities were compared with conventional static SENSE reconstructions. RESULTS The g-factor calculations showed that fast modulation of receive sensitivities in the order of the ADC dwell time during k-space acquisition can improve parallel imaging performance, as this effectively makes spatial information of both configurations simultaneously available for image encoding. This was confirmed by in vivo measurements, for which lower reconstruction errors (SSIM = 0.81 for acceleration R = 4) and g-factors (max g = 2.4; R = 4) were observed for the case of rapidly switched sensitivities compared to conventional reconstruction with static sensitivities (SSIM = 0.74 and max g = 3.2; R = 4). As the method relies on the short RF wavelength at ultra-high field, it does not yield significant benefits at 3T and below. CONCLUSIONS Time-varying receive sensitivities can be achieved by inserting PIN diodes in the receive loop coils, which allow modulation of B$_{1}$ $^{-}$ patterns. This offers an additional degree of freedom for image encoding, with the potential for improved parallel imaging performance at ultra-high field

In vivo characterization of the downfield part of 1 H MR spectra of human brain at 9.4 T: Magnetization exchange with water and relation to conventionally determined metabolite content

PURPOSE: To perform exchange-rate measurements on the in vivo human brain downfield spectrum (5-10 ppm) at 9.4 T and to compare the variation in concentrations of the downfield resonances and of known upfield metabolites to determine potential peak labels. METHODS: Non-water-suppressed metabolite cycling was used in combination with an inversion transfer technique in two brain locations in healthy volunteers to measure the exchange rates and T1 values of exchanging peaks. Spectra were fitted with a heuristic model of a series of 13 or 14 Voigt lines, and a Bloch-McConnell model was used to fit the exchange rate curves. Concentrations from non-water-inverted spectra upfield and downfield were compared. RESULTS: Mean T1 values ranged from 0.40 to 0.77 s, and exchange rates from 0.74 to 13.8 s-1 . There were no significant correlations between downfield and upfield concentrations, except for N-acetylaspartate, with a correlation coefficient of 0.63 and P < 0.01. CONCLUSIONS: Using ultrahigh field allowed improved separation of peaks in the 8.2 to 8.5 ppm amide proton region, and the exchange rates of multiple downfield resonances including the 5.8-ppm peak, previously tentatively assigned to urea, were measured in vivo in human brain. Downfield peaks consisted of overlapping components, and largely missing correlations between upfield and downfield resonances-although not conclusive-indicate limited contributions from metabolites present upfield to the downfield spectrum. Magn Reson Med, 2017. © 2017 International Society for Magnetic Resonance in Medicine

Paradoxical enhancement of atherosclerosis by probucol treatment in apolipoprotein E-deficient mice.

Dietary administration of probucol (0.5%, wt/wt) efficiently reduced total plasma cholesterol levels in apolipoprotein E-deficient mice (apoE-/-) by 40%, with decreases in high density lipoprotein (HDL) and apoAI by 70 and 50%, respectively. Paradoxically, however, aortic atherosclerotic plaques in the probucol-treated apoE-/- mice formed more rapidly than in the untreated apoE-/- mice, and the lesions were two to four times larger and more mature regardless of sex, age, and genetic background (P < 10(-)6). Histologically, lesions in probucol-treated mice contained increased fibrous materials and cells other than foam cells, and were commonly associated with focal inflammation and aneurysmal dilatation. Probucol treatment also accelerated lesion development in apoE+/- mice fed an atherogenic diet, indicating that the adverse effect is not dependent on the complete absence of apoE. Furthermore, mice lacking apoE and apoAI have plasma lipoprotein profiles very similar to the probucol-treated apoE-/- mice, but do not have accelerated plaque development. Thus, the enhanced atherosclerosis in the probucol-treated animals is unlikely to be caused by the reduction of HDL and apoAI levels. Our data indicate that a reduction in plasma cholesterol caused by probucol does not necessarily lead to an antiatherogenic effect

Clinical field-strength MRI of amyloid plaques induced by low-level cholesterol feeding in rabbits

Two significant barriers have limited the development of effective treatment of Alzheimer's disease. First, for many cases the aetiology is unknown and likely multi-factorial. Among these factors, hypercholesterolemia is a known risk predictor and has been linked to the formation of β-amyloid plaques, a pathological hallmark this disease. Second, standardized diagnostic tools are unable to definitively diagnose this disease prior to death; hence new diagnostic tools are urgently needed. Magnetic resonance imaging (MRI) using high field-strength scanners has shown promise for direct visualization of β-amyloid plaques, allowing in vivo longitudinal tracking of disease progression in mouse models. Here, we present a new rabbit model for studying the relationship between cholesterol and Alzheimer's disease development and new tools for direct visualization of β-amyloid plaques using clinical field-strength MRI. New Zealand white rabbits were fed either a low-level (0.125–0.25% w/w) cholesterol diet (n = 5) or normal chow (n = 4) for 27 months. High-resolution (66 × 66 × 100 µm3; scan time = 96 min) ex vivo MRI of brains was performed using a 3-Tesla (T) MR scanner interfaced with customized gradient and radiofrequency coils. β-Amyloid-42 immunostaining and Prussian blue iron staining were performed on brain sections and MR and histological images were manually registered. MRI revealed distinct signal voids throughout the brains of cholesterol-fed rabbits, whereas minimal voids were seen in control rabbit brains. These voids corresponded directly to small clusters of extracellular β-amyloid-positive plaques, which were consistently identified as iron-loaded (the presumed source of MR contrast). Plaques were typically located in the hippocampus, parahippocampal gyrus, striatum, hypothalamus and thalamus. Quantitative analysis of the number of histologically positive β-amyloid plaques (P < 0.0001) and MR-positive signal voids (P < 0.05) found in cholesterol-fed and control rabbit brains corroborated our qualitative observations. In conclusion, long-term, low-level cholesterol feeding was sufficient to promote the formation of extracellular β-amyloid plaque formation in rabbits, supporting the integral role of cholesterol in the aetiology of Alzheimer's disease. We also present the first evidence that MRI is capable of detecting iron-associated β-amyloid plaques in a rabbit model of Alzheimer's disease and have advanced the sensitivity of MRI for plaque detection to a new level, allowing clinical field-strength scanners to be employed. We believe extension of these technologies to an in vivo setting in rabbits is feasible and that our results support future work exploring the role of MRI as a leading imaging tool for this debilitating and life-threatening disease

Optimization of the Transmit and Receive Performance of the Transceiver Head Phased Array for Human Brain Imaging at Ultra-High Fields

Commonly, for optimal MRI performance at low (7T) magnetic fields (UHF) because large body coils are very inefficient at UHF, and cannot satisfy requirements in RF magnetic field, B1, anymore. To improve Tx-efficiency, B1/√P, and achieve a good Rx-performance, a local Transmit-Only volume coil (or Tx-array) in combination with a smaller tight-fit Receive-Only array, a ToRo coil, is used instead. Since most of the commercial UHF scanners provide only 8 kW of RF power, half of which is lost on a way to the coil, ToRo design still does not deliver sufficient B1. Tight-fit transceiver (TxRx) phased arrays improve Tx-efficiency in comparison to Tx-only arrays, which are larger to fit Rx-only arrays inside. However, the number of elements in a TxRx-array is restricted by the number of available RF Tx-channels (commonly <16) and difficulties associated with decoupling during transmission. In this work, we optimized the decoupling and developed new methods to increase the number of Rx-elements without compromising the Tx-performance. Thus, both the Tx- and Rx-performance of the TxRx-array can be optimized at same time

RF Arrays for Head & Body

Improvement of SNR is a critical step in designing any MRI RF coil. Use of arrays instead of a single coil is a major technique for the SNR enhancement. Designing an Rx-array commonly includes a choice of the type of elements (e.g. loops, striplines, dipoles), number of elements, decoupling method, coverage. The most common element of Rx-arrays is a surface loop, which has been used for human Rx-array designs at lower (1.5T, 3T) and ultra-high field (UHF, >7T). The presentation overviews most important steps of designing RF arrays including an optimization of the individual elements, decoupling, detuning, interfacing, cable routing

Transverse Electromagnetic (TEM) Surface Coils for Extremities

High‐field RF volume coils for the human limbs and heads are commonly formed from rigid cylinders. For extremity imaging this design often requires using larger one‐size‐fit‐all volume coils that decrease transmission efficiency and SNR. Splitting the coil solves this problem by providing tighter fit. It also improves patient access by eliminating the need for the coil to be slid over the region of interest. Split unshielded birdcage volume coils have been described previously for field strengths up to 3 T. For birdcage coils a direct electrical connection between two halves of the coil is required. For high‐field (>3 T) shielded birdcage coils, both the shield and the coil must be split and reliably connected electrically, which complicates the design. This problem can be circumvented by the use of split TEM volume coils. Since the elements of a TEM coil are coupled inductively, no direct electrical connection between the halves is necessary. In this study we demonstrate that the effects of splitting the shield for head and knee TEMs can be compensated for, and performance retained. For the knee, the improved access allowed the coil diameter to be reduced, enhancing the sensitivity by 15–20%