William M. Spees

Water proton MR properties of human blood at 1.5 Tesla: Magnetic susceptibility, T1, T2, T *2, and non‐Lorentzian signal behavior

Accurate knowledge of the magnetic properties of human blood is required for the precise modeling of functional and vascular flow‐related MRI. Herein are reported determinations of the relaxation parameters of blood, employing in vitro samples that are well representative of human blood in situ. The envelope of the blood 1H2O free‐induction decay signal magnitude during the first 100 msec following a spin echo at time TE is well‐ described empirically by an expression of the form, S(t) = So · exp{–R  *2 · (t – TE) – AR* · (t – TE)2}. The relaxation parameters AR* and R  *2 increase as a function of the square of the susceptibility difference between red blood cell and plasma and depend on the spin‐echo time. The Gaussian component, AR*, should be recognized in accurate modeling of MRI phenomena that depend upon the magnetic state of blood. The magnetic susceptibility difference between fully deoxygenated and fully oxygenated red blood cells at 37°C is 0.27 ppm, as determined independently by MR and superconducting quantum interference device (SQUID) measurements. This value agrees well with the 1936 report of Pauling and Coryell (Proc Natl Acad Sci USA 1936;22:210–216), but is substantially larger than that frequently used in MRI literature. Magn Reson Med 45:533–542, 2001.

A novel technology for the imaging of acidic prostate tumors by positron emission tomography.

Solid tumors often develop an acidic environment due to the Warburg effect. The effectiveness of diagnosis and therapy may therefore be enhanced by the design and use of pH-sensitive agents that target acidic tumors. Recently, a novel technology was introduced to target acidic tumors using pH low insertion peptide (pHLIP), a peptide that inserts across cell membranes as an alpha-helix when the extracellular pH (pH(e)) is acidic. In this study, we expanded the application of the pHLIP technology to include positron emission tomography imaging of the acidic environment in prostate tumors using (64)Cu conjugated to the pHLIP ((64)Cu-DOTA-pHLIP). Studies showed that this construct avidly accumulated in LNCaP and PC-3 tumors, with higher uptake and retention in the LNCaP tumors. Uptake correlated with differences in the bulk pH(e) of PC-3 and LNCaP tumors measured in magnetic resonance spectroscopy experiments by the (31)P chemical shift of the pH(e) marker 3-aminopropylphosphonate. This article introduces a novel class of noninvasive pH-selective positron emission tomography imaging agents and opens new research directions in the diagnosis of acidic solid tumors.

Pitx1 haploinsufficiency causes clubfoot in humans and a clubfoot-like phenotype in mice

Clubfoot affects 1 in 1000 live births, although little is known about its genetic or developmental basis. We recently identified a missense mutation in the PITX1 bicoid homeodomain transcription factor in a family with a spectrum of lower extremity abnormalities, including clubfoot. Because the E130K mutation reduced PITX1 activity, we hypothesized that PITX1 haploinsufficiency could also cause clubfoot. Using copy number analysis, we identified a 241 kb chromosome 5q31 microdeletion involving PITX1 in a patient with isolated familial clubfoot. The PITX1 deletion segregated with autosomal dominant clubfoot over three generations. To study the role of PITX1 haploinsufficiency in clubfoot pathogenesis, we began to breed Pitx1 knockout mice. Although Pitx1(+/-) mice were previously reported to be normal, clubfoot was observed in 20 of 225 Pitx1(+/-) mice, resulting in an 8.9% penetrance. Clubfoot was unilateral in 16 of the 20 affected Pitx1(+/-) mice, with the right and left limbs equally affected, in contrast to right-sided predominant hindlimb abnormalities previously noted with complete loss of Pitx1. Peroneal artery hypoplasia occurred in the clubfoot limb and corresponded spatially with small lateral muscle compartments. Tibial and fibular bone volumes were also reduced. Skeletal muscle gene expression was significantly reduced in Pitx1(-/-) E12.5 hindlimb buds compared with the wild-type, suggesting that muscle hypoplasia was due to abnormal early muscle development and not disuse atrophy. Our morphological data suggest that PITX1 haploinsufficiency may cause a developmental field defect preferentially affecting the lateral lower leg, a theory that accounts for similar findings in human clubfoot.

Imaging Intratumoral Convection: Pressure-Dependent Enhancement in Chemotherapeutic Delivery to Solid Tumors

Purpose: Low–molecular weight (LMW) chemotherapeutics are believed to reach tumors through diffusion across capillary beds as well as membrane transporters. Unexpectedly, the delivery of these agents seems to be augmented by reductions in tumor interstitial fluid pressure, an effect typically associated with high–molecular weight molecules that reach tumors principally through convection. We investigated the hypothesis that improved intratumoral convection can alter tumor metabolism and enhance the delivery of a LMW chemotherapeutic agent to solid tumors. Experimental Design: For this purpose, we applied 31P/19F magnetic resonance spectroscopy (MRS) and magnetic resonance spectroscopic imaging (MRSI) to examine the influence of type I collagenase on tumor bioenergetics and the delivery of 5-fluorouracil (5FU) to HT29 human colorectal tumors grown s.c. in mice. Results: Collagenase effected a 34% reduction in tumor interstitial fluid pressure with an attendant disintegration of intratumoral collagen. Neither mice-administered collagenase nor controls receiving PBS showed changes in 31phosphorus MRS–measured tumor bioenergetics; however, a time-dependent increase in the content of extracellular inorganic phosphate (Pie) was observed in tumors of collagenase-treated animals. 31Phosphorus MRSI showed that this increase underscored a more homogeneous distribution of Pie in tumors of experimental mice. 19Fluorine MRS showed that these changes were associated with a 50% increase in 5FU uptake in tumors of experimental versus control animals; however, this increase resulted in an increase in 5FU catabolites rather than fluoronucleotide intermediates that are required for subsequent cytotoxicity. Conclusions: These data indicate that the modulation of convective flow within tumors can improve the delivery of (LMW) chemotherapeutics and show the potential role for noninvasive imaging of this process in vivo.

Quantification and compensation of eddy-current-induced magnetic-field gradients

Two robust techniques for quantification and compensation of eddy-current-induced magnetic-field gradients and static magnetic-field shifts (ΔB0) in MRI systems are described. Purpose-built 1-D or six-point phantoms are employed. Both procedures involve measuring the effects of a prior magnetic-field-gradient test pulse on the phantoms free induction decay (FID). Phantom-specific analysis of the resulting FID data produces estimates of the time-dependent, eddy-current-induced magnetic field gradient(s) and ΔB0 shift. Using Bayesian methods, the time dependencies of the eddy-current-induced decays are modeled as sums of exponentially decaying components, each defined by an amplitude and time constant. These amplitudes and time constants are employed to adjust the scanners gradient pre-emphasis unit and eliminate undesirable eddy-current effects. Measurement with the six-point sample phantom allows for simultaneous, direct estimation of both on-axis and cross-term eddy-current-induced gradients. The two methods are demonstrated and validated on several MRI systems with actively-shielded gradient coil sets.

Imaging transgene activity in vivo.

The successful translation of gene therapy for clinical application will require the assessment of transgene activity as a measure of the biological function of a therapeutic transgene. Although current imaging permits the noninvasive detection of transgene expression, the critical need for quantitative imaging of the action of the expressed transgene has not been met. In vivo magnetic resonance spectroscopic imaging (MRSI) was applied to quantitatively delineate both the concentration and activity of a cytosine deaminase-uracil phosphoribosyltransferase (CD-UPRT) fusion enzyme expressed from a transgene. MRSI enabled the generation of anatomically accurate maps of the intratumoral heterogeneity in fusion enzyme activity. We observed an excellent association between the CD-UPRT concentration and activity and the percentage of CD-UPRT(+) cells. Moreover, the regional levels of UPRT activity, as measured by imaging, correlated well with the biological affect of the enzyme. This study presents a translational imaging paradigm for precise, in vivo measurements of transgene activity with potential applications in both preclinical and clinical settings.

White-matter diffusion fMRI of mouse optic nerve.

Non-invasive assessment of white-matter functionality in the nervous system would be a valuable basic neuroscience and clinical diagnostic tool. Using standard MRI techniques, a visual-stimulus-induced 27% decrease in the apparent diffusion coefficient of water perpendicular to the axonal fibers (ADC(perpendicular)) is demonstrated for C57BL/6 mouse optic nerve in vivo. No change in ADC(||) (diffusion parallel to the optic nerve fibers) was observed during visual stimulation. The stimulus-induced changes are completely reversible. A possible vascular contribution was sought by carrying out the ADC(perpendicular) measurements in hypercapnic mice with and without visual stimulus. Similar effects were seen in room-air-breathing and hypercapnic animals. The in vivo stimulus-induced ADC(perpendicular) decreases are roughly similar to literature reports for ex vivo rat optic nerve preparations under conditions of osmotic swelling. The experimental results strongly suggest that osmotic after-effects of nerve impulses through the axonal fibers are responsible for the observed ADC decrease.

Manganese‐enhanced MRI (MEMRI) via topical loading of Mn2+ significantly impairs mouse visual acuity: a comparison with intravitreal injection

Manganese‐enhanced MRI (MEMRI) with topical loading of MnCl2 provides optic nerve enhancement comparable to that seen by intravitreal injection. However, the impact of this novel and non‐invasive Mn2+ loading method on visual function requires further assessments. The objective of this study is to determine the optimal topical Mn2+ loading dosage for MEMRI and to assess visual function after MnCl2 loading. Intravitreal administration was performed to compare the two approaches of MnCl2 loading. Twenty‐four hours after topical loading of 0, 0.5, 0.75, and 1 M MnCl2, T1‐weighted, T2‐weighted, diffusion tensor imaging and visual acuity (VA) assessments were performed to determine the best topical loading dosage for MEMRI measurements and to assess the integrity of retinas and optic nerves. Mice were perfusion fixed immediately after in vivo experiments for hematoxylin and eosin and immunohistochemistry staining. Topical loading of 1 M MnCl2 damaged the retinal photoreceptor layer with no detectable damage to retina ganglion cell layers or prechiasmatic optic nerves. For the topical loading, 0.75 M MnCl2 was required to see sufficient enhancement of the optic nerve. At this concentration the visual function was significantly affected, followed by a slow recovery. Intravitreal injection (0.25 μL of 0.2 M MnCl2) slightly affected VA, with full recovery a day later. To conclude, intravitreal MnCl2injection provides more reproducible results with less adverse side‐effects than topical loading. Copyright

Use of ethylene glycol to evaluate gradient performance in gradient-intensive diffusion MR sequences

Imaging a phantom of known dimensions is a widely used and simple method for calibrating MRI gradient strength. However, full‐range characterization of gradient response is not achievable using this approach. Measurement of the apparent diffusion coefficient of a liquid with known diffusivity allows for calibration of gradient amplitudes across a wider dynamic range. An important caveat is that the temperature dependence of the liquids diffusion characteristics must be known, and the temperature of the calibration phantom must be recorded. In this report, we demonstrate that the diffusion coefficient of ethylene glycol is well described by Arrhenius‐type behavior across the typical range of ambient MRI magnet temperatures. Because of ethylene glycols utility as an NMR chemical‐shift thermometer, the same 1H MR spectroscopy measurements that are used for gradient calibration also simultaneously “report” the sample temperature. The high viscosity of ethylene glycol makes it well‐suited for assessing gradient performance in demanding diffusion‐weighted imaging and spectroscopy sequences. Magn Reson Med, 2012.

Diffusion fMRI detects white-matter dysfunction in mice with acute optic neuritis

Optic neuritis is a frequent and early symptom of multiple sclerosis (MS). Conventional magnetic resonance (MR) techniques provide means to assess multiple MS-related pathologies, including axonal injury, demyelination, and inflammation. A method to directly and non-invasively probe white-matter function could further elucidate the interplay of underlying pathologies and functional impairments. Previously, we demonstrated a significant 27% activation-associated decrease in the apparent diffusion coefficient of water perpendicular to the axonal fibers (ADC⊥) in normal C57BL/6 mouse optic nerve with visual stimulation using diffusion fMRI. Here we apply this approach to explore the relationship between visual acuity, optic nerve pathology, and diffusion fMRI in the experimental autoimmune encephalomyelitis (EAE) mouse model of optic neuritis. Visual stimulation produced a significant 25% (vs. baseline) ADC⊥ decrease in sham EAE optic nerves, while only a 7% (vs. baseline) ADC⊥ decrease was seen in EAE mice with acute optic neuritis. The reduced activation-associated ADC⊥ response correlated with post-MRI immunohistochemistry determined pathologies (including inflammation, demyelination, and axonal injury). The negative correlation between activation-associated ADC⊥ response and visual acuity was also found when pooling EAE-affected and sham groups under our experimental criteria. Results suggest that reduction in diffusion fMRI directly reflects impaired axonal-activation in EAE mice with optic neuritis. Diffusion fMRI holds promise for directly gauging in vivo white-matter dysfunction or therapeutic responses in MS patients.