Multi-parametric artificial neural network fitting of phase-cycled balanced steady-state free precession data.

PURPOSE: Standard relaxation time quantification using phase-cycled balanced steady-state free precession (bSSFP), eg, motion-insensitive rapid configuration relaxometry (MIRACLE), is subject to a considerable underestimation of tissue T1 and T2 due to asymmetric intra-voxel frequency distributions. In this work, an artificial neural network (ANN) fitting approach is proposed to simultaneously extract accurate reference relaxation times (T1 , T2 ) and robust field map estimates ( B1+ , ΔB0 ) from the bSSFP profile. METHODS: Whole-brain bSSFP data acquired at 3T were used for the training of a feedforward ANN with N = 12, 6, and 4 phase-cycles. The magnitude and phase of the Fourier transformed complex bSSFP frequency response served as input and the multi-parametric reference set [T1 , T2 , B1+ , ∆B0 ] as target. The ANN predicted relaxation times were validated against the target and MIRACLE. RESULTS: The ANN prediction of T1 and T2 for trained and untrained data agreed well with the reference, even for only four acquired phase-cycles. In contrast, relaxometry based on 4-point MIRACLE was prone to severe off-resonance-related artifacts. ANN predicted B1+ and ∆B0 maps showed the expected spatial inhomogeneity patterns in high agreement with the reference measurements for 12-point, 6-point, and 4-point bSSFP phase-cycling schemes. CONCLUSION: ANNs show promise to provide accurate brain tissue T1 and T2 values as well as reliable field map estimates. Moreover, the bSSFP acquisition can be accelerated by reducing the number of phase-cycles while still delivering robust T1 , T2 , B1+ , and ∆B0 estimates.

Expression changes of inflammatory cytokines TNF-α, IL-1ß and HO-1 in hematoma surrounding brain areas after intracerebral hemorrhage.

To study the expression changes of inflammatory factors heme oxygenase-1 (HO-1), tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß) in intracerebral hemorrhage (ICH), brain tissues surrounding hematoma were collected from ICH patients. The expressions of HO-1, TNF-α, IL- 1ß, and other genes were examined at different time points of ICH. Changes in HO-1, TNF-α, and IL-1ß positive cell numbers after ICH were detected by immunohistochemical staining. The results showed that the expressions of HO-1, TNF-α, and IL-1ß had no significant changes in brain tissues surrounding hematoma within 6 hours after ICH (P > 0.05). Their expressions during 6-24 hours and 24-72 hours after ICH increased constantly. After reaching the peak, they remained steady or slightly decreased after 72 hours. The dynamic expression changes of HO-1, TNF-α, and IL-1ß were observed and their development trends were interfered timely to alleviate the secondary neurological impairment after ICH, which was significant to prevent ICH.

Snapshot whole-brain T1 relaxometry using steady-state prepared spiral multislice variable flip angle imaging.

PURPOSE: Variable flip angle (VFA) imaging is widely used for whole-brain T1 quantification. Because of the requirement to acquire at least two sets of MR images at different flip angles, VFA relaxometry is relatively slow. Here, whole-brain VFA T1 mapping at 1.5 T is accelerated by using efficient spiral non-Cartesian imaging METHODS: To quantify T1 in the human brain, radiofrequency spoiled gradient-echo imaging is performed at two optimized flip angles using an interleaved 2D multislice sequence with high spoiling efficiency. The acquisitions are accelerated by using a spiral trajectory with 10 interleaves combined with a dedicated magnetization preparation to ensure steady-state conditions in minimal time. RESULTS: The investigated MR scan protocol allowed the acquisition of whole-brain T1 maps at a clinically relevant resolution in only 40 s (0.7 s per slice) with high reproducibility. White and gray matter T1 peaks clearly could be delineated by calculation of whole-brain T1 histograms, and the delivered T1 values showed good agreement with the reference method in selected regions of interest. CONCLUSION: Due to the use of a fast spiral k-space trajectory, whole-brain VFA T1 mapping could be accelerated by an order of magnitude compared to conventional Cartesian sampling. Magn Reson Med 79:856-866, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Rapid and robust variable flip angle T1 mapping using interleaved two-dimensional multislice spoiled gradient echo imaging.

PURPOSE: Conventional T1 mapping using three-dimensional (3D) radiofrequency (RF) spoiled gradient echo (SPGR) imaging with short repetition times (TR) is adversely affected by incomplete spoiling (i.e. residual T2 dependency). In this work, an optimized interleaved 2D multislice SPGR sequence scheme and an adapted postprocessing procedure are evaluated for highly T2 -insensitive T1 quantification of human brain tissues. METHODS: An efficient 2D multislice SPGR protocol including a relatively long TR of 200 ms is investigated with careful consideration of cross talk and magnetization transfer effects. Based on the derived scan protocol, T1 is quantified from the signal ratio of two SPGR datasets acquired at different flip angles. The effect of nonideal RF excitation profiles is incorporated into the SPGR signal model by performing Bloch simulations. RESULTS: Simulations showed that the parameters of the SPGR protocol (such as TR and the spoiler gradient moments) guarantee virtually complete spoiling. This result was confirmed by T1 measurements both in vitro using a 2% agar probe doped with 0.1 mM Gd (Gadovist) and in vivo in the human brain. CONCLUSION: The derived 2D multislice SPGR protocol offers efficient, highly reproducible, and in particular T2 -insensitive T1 quantification of human brain tissues. Magn Reson Med 77:1606-1611, 2017. © 2016 International Society for Magnetic Resonance in Medicine.

Variable flip angle T1 mapping in the human brain with reduced T2 sensitivity using fast radiofrequency-spoiled gradient echo imaging.

PURPOSE: Variable flip angle (VFA) T1 quantification using three-dimensional (3D) radiofrequency (RF) spoiled gradient echo imaging offers the acquisition of whole-brain T1 maps in clinically acceptable times. However, conventional VFA T1 relaxometry is biased by incomplete spoiling (i.e., residual T2 dependency). A new postprocessing approach is proposed to overcome this T2-related bias. METHODS: T1 is quantified from the signal ratio of two spoiled gradient echo (SPGR) images acquired at different flip angles using an analytical solution for the RF-spoiled steady-state signal in combination with a global T2 guess. T1 accuracy is evaluated from simulations and in vivo 3D SPGR imaging of the human brain at 3 Tesla. RESULTS: The simulations demonstrated that the sensitivity of VFA T1 mapping to T2 can considerably be reduced using a global T2 guess. The method proved to deliver reliable and accurate T1 values in vivo for white and gray matter in good agreement with inversion recovery reference measurements. CONCLUSION: Based on a global T2 estimate, the accuracy of VFA T1 relaxometry in the human brain can substantially be improved compared with conventional approaches which rely on the generally wrong assumption of ideal spoiling.

Optimization of measurement of mitochondrial electron transport activity in postmortem human brain samples and measurement of susceptibility to rotenone and 4-hydroxynonenal inhibition.

Mitochondrial function is required to meet the energetic and metabolic requirements of the brain. Abnormalities in mitochondrial function, due to genetic or developmental factors, mitochondrial toxins, aging or insufficient mitochondrial quality control contribute to neurological and psychiatric diseases. Studying bioenergetics from postmortem human tissues has been challenging due to the diverse range of human genetics, health conditions, sex, age, and postmortem interval. Furthermore, fresh tissues that were in the past required for assessment of mitochondrial respiratory function were rarely available. Recent studies established protocols to use in bioenergetic analyses from frozen tissues using animal models and cell cultures. In this study we optimized these methods to determine the activities of mitochondrial electron transport in postmortem human brain. Further we demonstrate how these samples can be used to assess the susceptibility to the mitochondrial toxin rotenone and exposure to the reactive lipid species 4-hydroxynonenal. The establishment of such an approach will significantly impact translational studies of human diseases by allowing measurement of mitochondrial function in human tissue repositories.

ATRX-DAXX Complex Expression Levels and Telomere Length in Normal Young and Elder Autopsy Human Brains.

The chromatin-remodeling complex ATRX/DAXX is one of the major epigenetic factors that controls heterochromatin maintenance due to its role in histone deposition. ATRX is involved in nucleosome configuration and maintenance of higher order chromatin structure, and DAXX is a specific histone chaperone for H3.3 deposition. Dysfunctions in this complex have been associated with telomere shortening, which influences cell senescence. However, data about this complex in brain tissue related to aging are still scarce. Therefore, in the present study, we analyzed ATRX and DAXX expressions in autopsied human brain specimens and the telomere length. A significant decrease in gene and protein expressions was observed in the brain tissues from the elderly compared with those from the young, which were related to short telomeres. These findings may motivate further functional analysis to confirm the ATRX-DAXX complex involvement in telomere maintenance and brain aging.

Multimodal imaging to explore endogenous fluorescence of fresh and fixed human healthy and tumor brain tissues.

To complement a project toward label-free optical biopsy and enhanced resection which the overall goal is to develop a multimodal nonlinear endomicroscope, this multimodal approach aims to enhance the accuracy in classifying brain tissue into solid tumor, infiltration and normal tissue intraoperatively. Multiple optical measurements based on one- and two-photon spectral and lifetime autofluorescence, including second harmonic generation imaging, were acquired. As a prerequisite, studying the effect of the time of measurement postexcision on tissue's spectral/lifetime fluorescence properties was warranted, so spectral and lifetime fluorescences of fresh brain tissues were measured using a point-based linear endoscope. Additionally, a comparative study on tissue's optical properties obtained by multimodal nonlinear optical imaging microscope from fresh and fixed tissue was necessary to test whether clinical validation of the nonlinear endomicroscope is feasible by extracting optical signatures from fixed tissue rather than from freshly excised samples. The former is generally chosen for convenience. Results of this study suggest that an hour is necessary postexcision to have consistent fluorescence intensities\lifetimes. The fresh (a,b,c) vs fixed (d,e,f) tissue study indicates that while all optical signals differ after fixation. The characteristic features extracted from one- and two-photon excitation still discriminate normal brain (a,d) cortical tissue, glioblastoma (GBM) (b,e) and metastases (c,f).

3D imaging in the postmortem human brain with CLARITY and CUBIC.

Recent innovations in tissue clearing and imaging technologies have enabled us to analyse biological systems directly in three-dimensions using thick samples. In this review, we discuss two of these recently reported tissue-clearing technologies (CLARITY and CUBIC) that are compatible with archival formalin-fixed human brain materials that have been fixed in formalin for a long period of time. We will discuss the pros and cons of these two technologies, examples of visualisation of Alzheimer neuropathological hallmarks and the exact protocols that we regularly use in the laboratory.

MECP2 Mutation Interrupts Nucleolin-mTOR-P70S6K Signaling in Rett Syndrome Patients.

Rett syndrome (RTT) is a severe and rare neurological disorder that is caused by mutations in the X-linked MECP2 (methyl CpG-binding protein 2) gene. MeCP2 protein is an important epigenetic factor in the brain and in neurons. In Mecp2-deficient neurons, nucleoli structures are compromised. Nucleoli are sites of active ribosomal RNA (rRNA) transcription and maturation, a process mainly controlled by nucleolin and mechanistic target of rapamycin (mTOR)-P70S6K signaling. Currently, it is unclear how nucleolin-rRNA-mTOR-P70S6K signaling from RTT cellular model systems translates into human RTT brain. Here, we studied the components of nucleolin-rRNA-mTOR-P70S6K signaling in the brain of RTT patients with common T158M and R255X mutations. Immunohistochemical examination of T158M brain showed disturbed nucleolin subcellular localization, which was absent in Mecp2-deficient homozygous male or heterozygote female mice, compared to wild type (WT). We confirmed by Western blot analysis that nucleolin protein levels are altered in RTT brain, but not in Mecp2-deficient mice. Further, we studied the expression of rRNA transcripts in Mecp2-deficient mice and RTT patients, as downstream molecules that are controlled by nucleolin. By data mining of published ChIP-seq studies, we showed MeCP2-binding at the multi-copy rRNA genes in the mouse brain, suggesting that rRNA might be a direct MeCP2 target gene. Additionally, we observed compromised mTOR-P70S6K signaling in the human RTT brain, a molecular pathway that is upstream of rRNA-nucleolin molecular conduits. RTT patients showed significantly higher phosphorylation of active mTORC1 or mTORC2 complexes compared to age- and sex-matched controls. Correlational analysis of mTORC1/2-P70S6K signaling pathway identified multiple points of deviation from the control tissues that may result in abnormal ribosome biogenesis in RTT brain. To our knowledge, this is the first report of deregulated nucleolin-rRNA-mTOR-P70S6K signaling in the human RTT brain. Our results provide important insight toward understanding the molecular properties of human RTT brain.

De novo identification and quantification of single amino-acid variants in human brain.

The detection of single amino-acid variants (SAVs) usually depends on single-nucleotide polymorphisms (SNPs) database. Here, we describe a novel method that discovers SAVs at proteome level independent of SNPs data. Using mass spectrometry-based de novo sequencing algorithm, peptide-candidates are identified and compared with theoretical protein database to generate SAVs under pairing strategy, which is followed by database re-searching to control false discovery rate. In human brain tissues, we can confidently identify known and novel protein variants with diverse origins. Combined with DNA/RNA sequencing, we verify SAVs derived from DNA mutations, RNA alternative splicing, and unknown post-transcriptional mechanisms. Furthermore, quantitative analysis in human brain tissues reveals several tissue-specific differential expressions of SAVs. This approach provides a novel access to high-throughput detection of protein variants, which may offer the potential for clinical biomarker discovery and mechanistic research.