Selective enrichment of endogenous peptides by chemically modified porous nanoparticles for peptidome analysis

We report the development of a combined strategy for high capacity, comprehensive enrichment of endogenous peptide from complex biological samples at natural pH condition. MCM-41 nanoparticles with highly ordered nanoscale pores (i.e. 4.8nm) and high-surface area (i.e. 751m(2)/g) were synthesized and modified with strong cation-exchange (SCX-MCM-41) and strong anion-exchange (SAX-MCM-41) groups. The modified nanoparticles demonstrated good size-exclusion effect for the adsorption of standard protein lysozyme with molecular weight (MW) of ca. 15kDa; and the peptides with MW lower than this value can be well adsorbed. Step elution of the enriched peptides with five salt concentrations presented that both modified nanoparticles have high capacity and complementarity for peptides enrichment, and the SAX-MCM-41 nanoparticles has obviously high selectivity for acidic peptides with pI (isoelectric point) lower than 4. Large-scale enrichment of endogenous peptides in 2mg mouse liver extract was achieved by further combination of SCX-MCM-41 and SAX-MCM-41 with unmodified MCM-41 nanoparticles. On-line 2D nano-LC/MS/MS was applied to analyze the enriched samples, and 2721 unique peptides were identified in total. Two-dimensional analysis of MW versus pI distribution combined with abundance of the identified peptides demonstrated that the three types of nanoparticles have comprehensive complementarity for peptidome enrichment.     

Development and characterization of rodent cardiac phantoms: comparison with in vivo cardiac imaging

The increasing availability of rodent models of human cardiovascular disease has led to a need to translate noninvasive imaging techniques such as magnetic resonance imaging (MRI) from the clinic to the animal laboratory. The aim of this study was to develop phantoms simulating the short-axis view of left ventricular motion of rats and mice, thus reducing the need for live animals in the development of MRI. Cylindrical phantoms were moulded from polyvinyl alcohol (PVA) Cryogel and attached via stiff water-filled tubing to a gear pump. Pulsatile distension of the phantoms was effected by suitable programming of the pump. Cine MRI scanning was carried out at 7 T and compared with in vivo rodent cardiac imaging. Suitable pulsatile performance was achieved with phantoms for which the PVA material had been subjected to two freeze-thaw cycles, resulting in T1 and T2 relaxation time constants of 1656±124 ms and 55±10 ms, respectively. For the rat phantom operating at 240 beats per min (bpm), the dynamic range of the outer diameter was from 10.3 to 12.4 mm with the wall thickness varying between 1.9 and 1.2 mm. Corresponding figures for the mouse phantom at 480 bpm were outer diameter range from 5.4 to 6.4 mm and wall thickness from 1.5 to 1.2 mm. Dynamic cardiac phantoms simulating rodent left ventricular motion in the short-axis view were successfully developed and compared with in vivo imaging. The phantoms can be used for future development work with reduced need of live animals.     

2D myocardial strain assessment in the mouse: a comparison between a synthetic lateral phase approach and block-matching using computer simulation

Estimating myocardial strain in the mouse with clinical equipment remains difficult due to the high heart rate and the small size of the mouse heart. Measuring the strain component perpendicular to the ultrasound beam is especially challenging because of the lack of phase information in that direction and the large speckle width compared to the wall thickness. In this study, the performance of a Synthetic Lateral Phase (SLP) approach was contrasted to a standard and a regularized 2D Speckle Tracking (2D ST) algorithm using simulated data sets. SLP yielded higher rms errors for the lateral strain estimates than the regularized 2D ST (Lateral rms error: 0.087 ± 0.012 vs. 0.052 ± 0.010; p < 0.05). No significant difference was found between the standard 2D ST and SLP. For the axial strain estimates, SLP produced higher rms errors than the standard 2D ST (Axial rms error: 0.063 ± 0.012 vs. 0.040 ± 0.008; p < 0.05). 2D ST combined with geometric regularization showed thus to be the most accurate method.     

New antitumor sesquiterpenoids from Santalum album of Indian origin

Three new campherenane-type (1, 4, 7) and three new santalane-type (9, 11, 12) sesquiterpenoids, and two aromatic glycosides (21, 22) together with 12 known metabolites including α,β-santalols (14, 18), (E)-α,β-santalals (15, 19), α,β-santaldiols (16, 20), α-santalenoic acid (17), and vanillic acid 4-O-neohesperidoside were isolated from Santalum album chips of Indian origin. The structures of the new compounds, including absolute configurations, were elucidated by 1D- and 2D-NMR spectroscopic and chemical methods. The antitumor promoting activity of these isolates along with several neolignans previously isolated from the same source was evaluated for both in vitro Epstein-Barr virus early antigen (EBV-EA) activation and in vivo two-stage carcinogenesis assays. Among them, compound 1 exhibited a potent inhibitory effect on EBV-EA activation, and also strongly suppressed two-stage carcinogenesis on mouse skin.     

Detection of T2 changes in an early mouse brain tumor

The aim of the study was to determine the effect of early tumor growth on T₂ relaxation times in an experimental glioma model. A 9.4-T magnetic resonance imaging (MRI) system was used for the investigations. An animal model (n=12) of glioma was established using an intracranial inoculation of U87MGdEGFRvIII cells. The imaging studies were performed from Day 10 through Day 13 following tumor inoculation. Tumor blood vessel density was determined using quantitative immunochemistry. Tumor volume was measured daily using MR images. T₂ values of the tumor were measured in five areas across the tumor and calculated using a single exponential fitting of the echo train. The measurements on Days 10 and 13 after tumor inoculation showed a 20% increase in T₂. The changes in T₂ correlated with the size of the tumor. Statistically significant differences in T₂ values were observed between the edge of the tumor and the brain tissue on Days 11, 12 and 13 (P=.014, .008, .001, respectively), but not on Day 10 (P=.364). The results show that T₂-weighted MRI may not detect glioma during an early phase of growth. T₂ increases in growing glioma and varies heterogenously across the tumor.     

Behavior of samarium inhaled by mice: Exposure length and time-dependent change

Distribution of samarium (Sm) in mice was investigated after inhalation exposure to Sm₂O₃ particles of 5 μm diameter and 15 mg/m³. Concentrations of Sm were determined by ICP-MS. Samarium was mainly distributed in lung, and the concentrations were varied, depending on the exposure length and time passed following final inhalation. Samarium concentrations in bone were lower than in lung, however, increased after cessation of the inhalation, suggesting that a part of Sm in lung was transported to bone via the blood stream.     

Comparison of mouse brain DTI maps using K-space average,image-space average,or no average approach

Diffusion tensor imaging (DTI) is achieved by collecting a series of diffusion-weighted images (DWIs). Signal averaging of multiple repetitions can be performed in the k-space (k-avg) or in the image space (m-avg) to improve the image quality. Alternatively, one can treat each acquisition as an independent image and use all of the data to reconstruct the DTI without doing any signal averaging (no-avg). To compare these three approaches, in this study, in vivo DTI data were collected from five normal mice. Noisy data with signal-to-noise ratios (SNR) that varied between five and 30 (before averaging) were then simulated. The DTI indices, including relative anisotropy (RA), trace of diffusion tensor (TR), axial diffusivity (λ║), and radial diffusivity (λ ⊥), derived from the k-avg, m-avg, and no-avg, were then compared in the corpus callosum white matter, cortex gray matter, and the ventricles. We found that k-avg and m-avg enhanced the SNR of DWI with no significant differences. However, k-avg produced lower RA in the white matter and higher RA in the gray matter, compared to the m-avg and no-avg, regardless of SNR. The latter two produced similar DTI quantifications. We concluded that k-avg is less preferred for DTI brain imaging.     

Relaxation dispersion in MRI induced by fictitious magnetic fields

A new method entitled Relaxation Along a Fictitious Field (RAFF) was recently introduced for investigating relaxations in rotating frames of rank ≥ 2. RAFF generates a fictitious field (E) by applying frequency-swept pulses with sine and cosine amplitude and frequency modulation operating in a sub-adiabatic regime. In the present work, MRI contrast is created by varying the orientation of E, i.e. the angle ε between E and the z″ axis of the second rotating frame. When ε > 45°, the amplitude of the fictitious field E generated during RAFF is significantly larger than the RF field amplitude used for transmitting the sine/cosine pulses. Relaxation during RAFF was investigated using an invariant-trajectory approach and the Bloch-McConnell formalism. Dipole-dipole interactions between identical (like) spins and anisochronous exchange (e.g., exchange between spins with different chemical shifts) in the fast exchange regime were considered. Experimental verifications were performed in vivo in human and mouse brain. Theoretical and experimental results demonstrated that changes in ε induced a dispersion of the relaxation rate constants. The fastest relaxation was achieved at ε ≈ 56°, where the averaged contributions from transverse components during the pulse are maximal and the contribution from longitudinal components are minimal. RAFF relaxation dispersion was compared with the relaxation dispersion achieved with off-resonance spin lock T(?ρ) experiments. As compared with the off-resonance spin lock T(?ρ) method, a slower rotating frame relaxation rate was observed with RAFF, which under certain experimental conditions is desirable.     

5’ and 3’ splicing signals evolution in vertebrates: Analysis in a conserved gene family

The mitochondrial solute carrier genes (SLC25) are highly conserved during vertebrate evolution. In most SLC25 genes of zebrafish, chicken, mouse, and human, the introns are located at exactly superimposable positions. In these topographically corresponding introns we studied the composition of the initial and terminal hexanucleotides (5’ss and 3’ss) which are instrumental in splicing signaling, focusing on the evolutionary conservation/mutation dynamics of these genetically related sequences. At each position, the per cent conservation of zebrafish individual nucleotides in chicken, mouse and human is proportional to their percent frequency in zebrafish; furthermore, nucleotide mutations are biased in favor of the more represented nucleotides, thus compensating for those highly represented zebrafish nucleotides which have not been conserved. As a result of these evolutionary dynamics, the general nucleotide composition at each position has remained relatively conserved throughout vertebrates. At 5’ss, following the canonical GT, A and G are largely prevailing at position +3, A at +4 and G at +5 (GT[A/G]AGx). At 3’ss, T and C are largely prevailing at positions −6, −5 and −3, preceding the canonical intron terminal AG ([C/T] [C/T]x[C/T]AG). However, the actual composition of the tetranucleotides at 5’ and 3’ often does not conform to the above scheme. At 5’ss the more canonical sequence is completely expressed in 63% of cases and partially (2 or 1 matches) in 37 % of cases. At 3’ss the more canonical sequence is completely expressed in 71 % of cases and partially (2 or 1 matches) in 29 % of cases. The nucleotide conservation loss (nucleotide mutation) is higher in the evolution from fish to the last common ancestor of birds and mammals (58 %), then diminishes in the successive evolution steps up to the mammalian common ancestor (10 %), and becomes still lower at the divergence of rodents and primates (5 %).     

Ablation of subsurface tumors using an ultra-short pulse laser

This paper demonstrates the novel technique of ablating subsurface tumors with minimal thermal damage to surrounding healthy tissue using a focused laser beam from an ultra-short pulse diode laser source. Experiments were performed on anesthetized healthy mice as well as mice with mammary tumors in order to demonstrate the fundamental advantages of using a focused-beam, ultra-short pulse laser to ablate subcutaneous tissues. The technique was demonstrated through histological analysis of tissue samples after irradiation of anesthetized mice with or without mammary tumors. To demonstrate the efficacy of subsurface focusing, temperature was monitored at the subsurface tumor location and at the surface in an untranslated sample while irradiating with a focused ultra-short pulsed 1552 nm laser. Results show that temperature rise was dramatically greater at the focal depth than at the surface. Irradiation at the subsurface tumor location while translated over time across the tumor location resulted in precise ablation of the tumor. This work shows that a focused-beam ultra-short pulse 1552 nm laser results in precise ablation at the desired location with high efficacy and a minimal zone of collateral thermal and/or mechanical damage.