Recruitment of beneficial M2 macrophages to injured spinal cord is orchestrated by remote brain choroid plexus.

Monocyte-derived macrophages are essential for recovery after spinal cord injury, but their homing mechanism is poorly understood. Here, we show that although of common origin, the homing of proinflammatory (M1) and the "alternatively activated" anti-inflammatory (M2) macrophages to traumatized spinal cord (SC) was distinctly regulated, neither being through breached blood-brain barrier. The M1 macrophages (Ly6c(hi)CX3CR1(lo)) derived from monocytes homed in a CCL2 chemokine-dependent manner through the adjacent SC leptomeninges. The resolving M2 macrophages (Ly6c(lo)CX3CR1(hi)) derived from monocytes trafficked through a remote blood-cerebrospinal-fluid (CSF) barrier, the brain-ventricular choroid plexus (CP), via VCAM-1-VLA-4 adhesion molecules and epithelial CD73 enzyme for extravasation and epithelial transmigration. Blockage of these determinants, or mechanical CSF flow obstruction, inhibited M2 macrophage recruitment and impaired motor-function recovery. The CP, along with the CSF and the central canal, provided an anti-inflammatory supporting milieu, potentially priming the trafficking monocytes. Overall, our finding demonstrates that the route of monocyte entry to central nervous system provides an instructional environment to shape their function.

Major mouse placental compartments revealed by diffusion-weighted MRI, contrast-enhanced MRI, and fluorescence imaging.

Mammalian models, and mouse studies in particular, play a central role in our understanding of placental development. Magnetic resonance imaging (MRI) could be a valuable tool to further these studies, providing both structural and functional information. As fluid dynamics throughout the placenta are driven by a variety of flow and diffusion processes, diffusion-weighted MRI could enhance our understanding of the exchange properties of maternal and fetal blood pools--and thereby of placental function. These studies, however, have so far been hindered by the small sizes, the unavoidable motions, and the challenging air/water/fat heterogeneities, associated with mouse placental environments. The present study demonstrates that emerging methods based on the spatiotemporal encoding (SPEN) of the MRI information can robustly overcome these obstacles. Using SPEN MRI in combination with albumin-based contrast agents, we analyzed the diffusion behavior of developing placentas in a cohort of mice. These studies successfully discriminated the maternal from the fetal blood flows; the two orders of magnitude differences measured in these fluids' apparent diffusion coefficients suggest a nearly free diffusion behavior for the former and a strong flow-based component for the latter. An intermediate behavior was observed by these methods for a third compartment that, based on maternal albumin endocytosis, was associated with trophoblastic cells in the interphase labyrinth. Structural features associated with these dynamic measurements were consistent with independent intravital and ex vivo fluorescence microscopy studies and are discussed within the context of the anatomy of developing mouse placentas.

Boron analysis and boron imaging in biological materials for Boron Neutron Capture Therapy (BNCT).

Boron Neutron Capture Therapy (BNCT) is based on the ability of the stable isotope 10B to capture neutrons, which leads to a nuclear reaction producing an alpha- and a 7Li-particle, both having a high biological effectiveness and a very short range in tissue, being limited to approximately one cell diameter. This opens the possibility for a highly selective cancer therapy. BNCT strongly depends on the selective uptake of 10B in tumor cells and on its distribution inside the cells. The chemical properties of boron and the need to discriminate different isotopes make the investigation of the concentration and distribution of 10B a challenging task. The most advanced techniques to measure and image boron are described, both invasive and non-invasive. The most promising approach for further investigation will be the complementary use of the different techniques to obtain the information that is mandatory for the future of this innovative treatment modality.

Quantification of Mouse Renal Perfusion Using Arterial Spin Labeled MRI at 1 T.

RATIONALE AND OBJECTIVES: Quantitative measurement of renal perfusion in murine models provides important information on the organ physiology and disease states. The 1-T desktop magnetic resonance imaging has a small footprint and a self-contained fringe field. This resultant flexibility in siting makes the system ideal for preclinical imaging research. Our objective was to evaluate the capability of the 1-T desktop magnetic resonance imaging to measure mouse renal perfusion without the administration of exogenous contrast agents. MATERIALS AND METHODS: We implemented a flow-sensitive alternating inversion recovery (FAIR)-based arterial spin labeling sequence with a mouse volume coil on a 1-T desktop magnetic resonance scanner. The validity of the implementation was tested by comparing obtained renal perfusion results with literature values for normal mice and challenging the technique with mice treated with furosemide, a blood vessel vasoconstrictor drug. RESULTS: The measured cortical and medullary perfusions were quantified to be 402 ± 95 and 184 ± 52 mL/100 g/min, respectively, in agreement with literature values. The ratio of cortical to medullary renal blood flow was between 2 and 3 and was independent of the mouse weight. As expected, upon furosemide injection, a decrease (~50%) in cortical perfusion was observed in the mice population, at 1 hour post injection compared to baseline (P < 0.0001), which returned to baseline after 24 hours (P = 0.68). CONCLUSIONS: We reported the successful application of FAIR-based arterial spin labeling for noncontrast perfusion measurement of mouse kidneys using a 1-T desktop scanner. The easy implementation of FAIR sequence on a 1-T desktop scanner offers the potential for longitudinal perfusion studies in limited access areas such as behind the barrier in mouse facilities and in multimodality preclinical imaging laboratories without the administration of exogenous contrast agents.

Noninvasive quantitative in vivo mapping and metabolism of boronophenylalanine (BPA) by nuclear magnetic resonance (NMR) spectroscopy and imaging.

10B-enriched L-p-boronophenylalanine (BPA) is one of the compounds used in boron neutron capture therapy (BNCT). In this study, several variations of nuclear magnetic resonance spectroscopy (MRS) and spectroscopic imaging (MRSI) were applied to investigate the uptake, clearance and metabolism of the BPA-fructose complex (BPA-F) in normal mouse kidneys, rat oligodendroglioma xenografts, and rat blood. Localized 1H MRS was capable of following the uptake and clearance of BPA-F in mouse kidneys with temporal resolution of a few minutes, while 1H MRSI was used to image the BPA distribution in the kidney with a spatial resolution of 9 mm3. The results also revealed significant dissociation of the BPA-F complex to free BPA. This finding was corroborated by 1H and 11B NMR spectroscopy of rat blood samples as well as of tumor samples excised from mice after i.v. injection of BPA-F. This investigation demonstrates the feasibility of using 1H MRS and MRSI to follow the distribution of BPA in vivo, using NMR techniques specifically designed to optimize BPA detection. The implementation of such procedures could significantly improve the clinical efficacy of BNCT.

Electric field driven flow in natural porous media.

Electric fields were applied to fluid-saturated packed sand beds (0.23+/-0.03 mm average pore diameter), and the effects on the mobility of the water molecules were monitored using stimulated echo (STE) and pulsed field gradient (PFG) experiments. The mean flow velocity, averaged over the entire sample, is expected to vanish in closed systems, but the PFG and time dependent signal decay was enhanced beyond the effects of thermal diffusion, due to velocity dispersion. The internal flow generated by the electric field was shown to be fully time-reversible upon inverting the electric field polarity (for total flow times of up to 0.4s), a strong indication that the NMR detected displacements were mainly due to electro-osmotic flow (EOF). However, a comparison of the velocity dispersion for different electrolyte concentrations showed that the measured effect scaled with the applied power VI (V = voltage, I = electric current), rather than with the voltage alone, contrary to the prediction of the basic model for EOF in a single capillary channel.

A method for fat suppression in MRI based on diffusion-weighted imaging.

The diffusion coefficient of lipid molecules is usually much smaller than that of water, and it is demonstrated here how this difference can be exploited for robust fat suppression in magnetic resonance imaging (MRI). In contrast to the prevailing methods, diffusion-based fat suppression does not rely on chemical shift differences between water and lipids and can therefore be applied easily in low or inhomogeneous magnetic fields. It is also independent of relaxation times and can therefore be incorporated in experiments requiring conventional T(1)-weighted contrast. Diffusion-based fat suppression (DIFFSUP) consists of subtracting the signals acquired at low and high b-values, where the high b-value is ideally designed to achieve full suppression of the water and negligible attenuation of the lipid signal. Since high b-value images may be particularly affected by motion artifacts, a version of DIFFSUP incorporating first-order velocity compensation is also proposed and demonstrated. Results from phantoms and live mice at field strengths of 4.7 T and 1.0 T are presented.

Metabolism of borono-phenylalanine-fructose complex (BPA-fr) and borocaptate sodium (BSH) in cancer patients--results from EORTC trial 11001.

Within the clinical trial EORTC 11001, patients were infused with (10)B-enriched borono-phenylalanine-fructose complex (BPA-fr), or borocaptate sodium (BSH) solutions, which are used as boron carriers for boron neutron capture therapy. Urine samples were periodically collected and analyzed by (10)B NMR spectroscopy. The results revealed time-dependent metabolic changes of the administered compounds. BPA-fr dissociated to the constituents BPA and fructose, and the borate group was partly cleaved from BPA. BSH was partly aggregated to a dimer form, BSSB. These observations were previously reported for cultured cells and animal models, and are confirmed here in human cancer patients.

Quantification of slow flow using FAIR.

Phase contrast (PC)-based MRI methods are considered to be the most accurate approach for spatially resolved flow quantification, but the measurement of very slow velocities requires signal detection at long echo times and the application of strong field gradients. On the other hand, measurements based on time-of-flight or inflow effects can be conducted at short echo times and without flow-encoding gradients. A method for imaging flow at velocities of the order of 0.1 mm/s is presented and validated here. It consists of measuring the apparent spin-lattice relation rate (R(1)*) of the flowing fluid using magnetization preparation by alternating slice-selective and nonselective inversion pulses (FAIR or flow-sensitive alternating inversion recovery) and a fast gradient-echo detection sequence. This method is appropriate for the quantitative imaging of slow flow at low Reynolds numbers in fluids where the T(2) values are too short to allow sensitive flow measurements by phase contrast-based methods.

Biomedical applications of 10B and 11B NMR.

This review focuses mainly on the detection and investigation of molecules used for boron neutron capture therapy (BNCT) by 10B and 11B NMR. In this binary radiation treatment, boron-containing molecules (also called 'BNCT agents') enriched in the 10B isotope, are targeted to the tumor, and irradiated with thermal or epithermal neutrons. Capture of these neutrons by 10B nuclei generates cell-damaging radiation, confined to single cell dimensions. NMR research efforts have primarily been applied in two directions: first, to investigate the metabolism and pharmaco-kinetics of BNCT agents in-vivo, and second, to use localized NMR spectroscopy and/or MRI for non-invasive mapping of the administered molecules in treated animals or patients. While the first goal can be pursued using 11B NMR for natural-abundance samples (80% 11B / 20% 10B), molecules used in the actual treatment are > 95% enriched in 10B, and must therefore be detected by 10B NMR. Both 10B (spin 3) and 11B (spin 3/2) are quadrupolar nuclei, and their typical relaxation times, in common BNCT agents in biological environments, are rather short. This poses some technical challenges, particularly for MRI, which will be reviewed, along with possible solutions. The first attempts at 11B NMR and MRI detection of BNCT agents in biological tissue were conducted over a decade ago. Since then, results from 11B MRI in laboratory animals and in humans have been reported, and 11B NMR spectroscopy provided interesting and unique information about the metabolism of some BNCT agents in cultured cells. 10B NMR was applied either 'indirectly' (in double-resonance experiments involving coupled protons), but also by direct 10B MRI in mice. However, no results involving the NMR detection of 10B-enriched compounds in treated patients have been reported yet.

Optimized 1H MRS and MRSI methods for the in vivo detection of boronophenylalanine.

Boronophenylalanine (BPA) is used as Boron-10 carrier in boron neutron capture therapy, an experimental cancer radiotherapy. Results of quantitative, noninvasive in vivo detection and imaging of BPA in laboratory animals using (1)H NMR are presented for the first time. The purpose of this study was to implement and validate optimized techniques for the efficient detection of BPA. The (1)H NMR signals through which BPA is most readily detected in vivo are those from the aromatic ring of the molecule, which are part of a scalar-coupled spin system. The preferred detection method should therefore be based on a pulse sequence in which the effective TE is as short as possible. Modified versions of LASER (tau(CP) = 4.6 ms, TE = 27.6 ms) and double-echo slice-selective 2D MRSI (TE = 12 ms) were implemented for single-voxel spectroscopy and spectroscopic imaging of BPA, respectively. Chemical shift selective excitation was used for both sequences, based on a pulse that enabled narrow-band excitation without concomitant delay in TE. SI data without water suppression was used for absolute quantitation and for correction of B(0) variations. Experiments were conducted at 4.7 T in phantoms and in mice where the infused BPA was detected in the kidney.

Water diffusion in the different microenvironments of breast cancer.

The parameters that characterize the intricate water diffusion in tumors may serve to reveal their distinct pathology. Specifically, the application of diffusion magnetic resonance imaging (MRI) can aid in characterizing breast cancer, as well as monitoring response to therapy. We present here a non-invasive, quantitative MRI investigation, at high spatial resolution, of water diffusion in hormonal dependent MCF7 breast tumors implanted orthotopically in immunodeficient mice. Distinctive MRI protocols were designed in this study, utilizing a broad range of diffusion times and diffusion gradient strengths. Application of these protocols allowed water diffusion in the tissue extracellular and intracellular compartments to be distinguished, and the effect of restricted diffusion and water exchange on the water diffusion in these compartments to be evaluated. Pixel-by-pixel analysis yielded parametric maps of the estimated volume fraction and apparent diffusion coefficient of each compartment. The diffusion of the water in the extracellular microenvironment was approximately two fold slower than that of free water, and in the intracellular compartment was about one order of magnitude slower than that of free water and demonstrated restriction of water diffusion at long diffusion times. Mapping of the water fraction in each compartment was further employed to monitor changes during tumor progression and to assess tumor response to hormonal manipulation with a new antiestrogenic drug, tamoxifen methiodide (TMI). It was found that, in parallel to the growth arrest by this drug, the volume fraction of the slowly diffusing water increased, suggesting a TMI-induced cell swelling. This study can serve as a basis for extending diffusion breast MRI in the clinical setting.

Photodynamic therapy with Pd-Bacteriopheophorbide (TOOKAD): successful in vivo treatment of human prostatic small cell carcinoma xenografts.

Small cell carcinoma of the prostate (SCCP), although relatively rare, is the most aggressive variant of prostate cancer, currently with no successful treatment. It was therefore tempting to evaluate the response of this violent malignancy and its bone lesions to Pd-Bacteriopheophorbide (TOOKAD)-based photodynamic therapy (PDT), already proven by us to efficiently eradicate other aggressive non-epithelial solid tumors. TOOKAD is a novel bacteriochlorophyll-derived, second-generation photosensitizer recently, developed by us for the treatment of bulky tumors. This photosensitizer is endowed with strong light absorbance (epsilon(0) approximately 10(5) mol(-1) cm(-1)) in the near infrared region (lambda=763nm), allowing deep tissue penetration. The TOOKAD-PDT protocol targets the tumor vasculature leading to inflammation, hypoxia, necrosis and tumor eradication. The sensitizer clears rapidly from the circulation within a few hours and does not accumulate in tissues, which is compatible with the treatment of localized tumor and isolated metastases. Briefly, male CD1-nude mice were grafted with the human SCCP (WISH-PC2) in 3 relevant anatomic locations: subcutaneous (representing tumor mass), intraosseous (representing bone metastases) and orthotopically within the murine prostate microenvironment. The PDT protocol consisted of i.v. administration of TOOKAD (4 mg/kg), followed by immediate illumination (650-800 nm) from a xenon light source or a diode laser emitting at 770 nm. Controls included untreated animals or animals treated with light or TOOKAD alone. Tumor volume, human plasma chromogranin A levels, animal well being and survival were used as end points. In addition, histopathology and immunohistochemistry were used to define the tumor response. Subcutaneous tumors exhibited complete healing within 28-40 days, reaching an overall long-term cure rate of 69%, followed for 90 days after PDT. Intratibial WISH-PC2 lesions responded with complete tumor elimination in 50% of the treated mice at 70-90 days after PDT as documented histologically. The response of the orthotopic model was also analyzed histologically with similar results. The study with this model suggests that TOOKAD-based PDT can reach large tumors and is a feasible, efficient and well-tolerated approach for minimally invasive treatment of local and disseminated SCCP.