Thian C. Ng

NMR study of in vivo RIF-1 tumors. Analysis of perchloric acid extracts and identification of 1H, 31P and 13C resonances

Perchloric acid extracts of radiation-induced fibrosarcoma (RIF-1) tumors grown in mice have been analyzed by multinuclear NMR spectroscopy and by various chromatographic methods. This analysis has permitted the unambiguous assignment of the 31P resonances observed in vivo to specific phosphorus-containing metabolites. The region of the in vivo spectra generally assigned to sugar phosphates has been found in RIF-1 tumors to contain primarily phosphorylethanolamine and phosphorylcholine rather than glycolytic intermediates. Phosphocreatine was observed in extracts of these tumor cells grown in culture as well as in the in vivo spectra, indicating that at least some of the phosphocreatine observed in vivo arises from the tumor itself and not from normal tissues. In the 31P-NMR spectra of the perchloric acid extract, resonances originating from purine and pyrimidine nucleoside di- and triphosphate were resolved. HPLC analyses of the nucleotide pool indicate that adenine derivatives were the most abundant components, but other nucleotides were present in significant amounts. The 1H and 13C resonance assignments of the majority of metabolites present in RIF-1 extracts have also been made. Of particular importance is the ability to observe lactate, the levels of which may provide a noninvasive measure of glycolysis in these cells in both the in vitro states. In addition, the aminosulfonic acid, taurine, was found in high levels in the tumor extracts.

31P NMR spectroscopy of in vivo tumors

Abstract A probe, suitable for any wide-bore NMR spectrometer, was constructed for monitoring high-resolution spectra of in vivo subcutaneously implanted tumors in mice. Preliminary studies of a variety of murine tumors (MOPC 104E myeloma, Dunn osteosarcoma, colon-26, ovarian M5, and mammary adenocarcinoma as well as human colon, mammary, and lung tumors in athymic mice) indicate that the 31P NMR spectrum is a sensitive monitor of progressive metabolic changes that occur during untreated tumor growth and an early indicator of tumor response to chemotherapy, hyperthermia, and X radiation. Response to each of these therapeutic modalities is accompanied by distinctly different spectral changes.

Postictal Alteration of Sodium Content and Apparent Diffusion Coefficient in Epileptic Rat Brain Induced by Kainic Acid

Summary: Purpose: We studied temporal changes of brain sodium and apparent diffusion coefficient (ADC) in a temporal lobe epilepsy (TLE) rat model using kainic acid (KA).

1H-MRS evidence of neurodegeneration and excess glutamate glutamine in ALS medulla

Objective: To determine whether short echo-time (TE) proton magnetic resonance spectroscopic imaging (1H-MRSI) can detect in vivo differences in signal intensities of specific metabolites in the medulla of patients with ALS compared with healthy individuals and whether these metabolites could be useful surrogate markers of disease. Background: 1H-MRSI can detect N-acetylaspartate +N-acetylaspartylglutamate (abbreviated NAx), which is localized to neurons, and glutamate (Glu) + glutamine (Gln), abbreviated Glx, which may be important in ALS pathogenesis. The medulla is an ideal region to study ALS because of its high density of nuclei and fiber tracts that frequently undergo degeneration, even when more rostral brain regions show minimal pathology. Methods: Ten patients with ALS and seven healthy control subjects underwent short TE 1H-MRSI on a 1.5 T clinical imaging system. Signal intensities of NAx and Glx were normalized to creatine–phosphocreatine and compared between groups. Results: Compared with normal subjects, the medulla of patients with ALS had 17% lower NAx (p = 0.03) and 55% higher Glx (p = 0.02) signals. Bulbar symptoms, represented by the ALS Functional Rating Scale, correlated with Glx (r = −0.68, p = 0.03) but not NAx (r = 0.22, p = 0.53). Conclusion: There is in vivo 1H-MRSI evidence of neuronal degeneration or loss and excess Glu + Gln in the medulla of patients with ALS. Although this cross-sectional study cannot identify which change occurred first, the higher Glx signal in the medulla of patients with more dysarthria and dysphagia is consistent with the hypothesis of Glu excitotoxicity in ALS pathogenesis. Longitudinal 1H-MRSI studies of the medulla (and other brain regions) in more patients with ALS are required to confirm these findings and to determine whether such metabolite changes will be useful in monitoring disease progression, in clinical diagnosis, and in understanding the pathogenesis of ALS.

MRS Metabolic Markers of Seizures and Seizure‐Induced Neuronal Damage

Summary: Purpose: Proton magnetic resonance spectroscopy (MRS) was used to identify specific in situ metabolic markers for seizures and seizure‐induced neuronal damage. Kainic acid (KA)‐induced seizures lead to histopathologic changes in rat brain. The protective effect of cycloheximide treatment against neuronal damage caused by KA‐induced seizures was studied, using in situ proton MRS imaging technique.

Brain Tissue Sodium Is a Ticking Clock Telling Time After Arterial Occlusion in Rat Focal Cerebral Ischemia

BACKGROUND AND PURPOSE Many patients with acute stroke are excluded from receiving thrombolysis agents within the necessary time limit (3 or 6 hours from stroke onset) because they or their family members are unable provide the time of stroke onset. Brain tissue sodium concentration ([Na(+)]) increases gradually and incessantly during the initial hours of experimental focal cerebral ischemia but only in severely damaged brain regions. We propose that this steady increase in [Na(+)] can be used to estimate the time after arterial occlusion in the rat middle cerebral artery occlusion model of ischemic stroke. METHODS Sixteen anesthetized Sprague-Dawley rats underwent permanent middle cerebral artery occlusion combined with bilateral common artery occlusion. After 100 to 450 minutes, diffusion-weighted MRI was used to generate apparent diffusion coefficient (ADC) maps, cerebral blood flow (CBF) was determined with (14)C-iodoantipyrine (in a subset of 7 animals), and the brain was frozen. Autoradiographic CBF sections and punch samples for Na(+) analysis were obtained from the brain at the same level of the MR image. Severely at risk regions were identified with an ADC of <520 microm(2)/s and, in the subset, with both ADC of <520 microm(2)/s and CBF of <40 mL. 100 g(-1). min(-1). RESULTS Both CBF and the ADC dropped quickly and remained stable in the initial hours after ischemic onset. Linear regression revealed strong linearity between [Na(+)] and time after onset, with a slope of 0.95 or 1.00 (mEq/kg DW)/min, with both ADC and ADC-plus-CBF criteria, respectively. The 95% CIs at 180 and 360 minutes were between 41 and 52 minutes. CONCLUSIONS The time after ischemic onset can be estimated with this 2-step process. First, ADC and CBF are used to identify severely endangered regions. Second, the [Na(+)] in these regions is used to estimate time after onset. The favorable 95% CIs at the time limits for thrombolytic therapy and the availability of measurements of ADC, CBF, and [Na(+)] in humans through the use of MRI suggest that this time-estimation scheme could be used to assess the appropriateness of thrombolysis for patients who do not know when the stroke occurred.

Brain activation during human finger extension and flexion movements

Corticospinal projections to the motor neuron pool of upper-limb extensor muscles have been reported to differ from those of the flexor muscles in humans and other primates. The influence of this difference on the central nervous system control for extension and flexion movements is unknown. Cortical activation during thumb extension and flexion movements of eight human volunteers was measured using functional magnetic resonance imaging (fMRI), which detects signal changes caused by an alteration in the local blood oxygenation level. Although the relative activity of the extensor and flexor muscles of the thumb was similar, the brain volume activated during extension was substantially larger than that during flexion. These fMRI results were confirmed by measurements of EEG-derived movement-related cortical potential. Higher brain activity during thumb extension movement may be a result of differential corticospinal, and possibly other pathway projections to the motoneuron pools of extensor and flexor muscles of upper the extremities.

Human tumors as examined by in vivo 31P NMR in athymic mice

Abstract Surface coils have been employed to monitor in vivo 31P NMR spectra of human breast (MX-1), lung (LX-1) and colon (CX-1) tumors subcutaneously implanted in athymic mice. Spectra of these NCI screening tumors were measured during various stages of untreated growth. A progressive decrease in phosphocreatine and ATP and an increase in inorganic phosphate were observed for the MX-1 tumor line. The lung and colon tumors also exhibited a growth associated increase in sugar phosphates. The spectral characteristics of these tumors were very similar to those of corresponding murine tumors: mammary 16 C adenocarcinoma, Lewis lung sarcoma and colon 26. These data provide a basis for development of NMR techniques for monitoring human tumors in situ or in athymic mice.

Sensitivity of Magnetic Resonance Diffusion-Weighted Imaging and Regional Relationship Between the Apparent Diffusion Coefficient and Cerebral Blood Flow in Rat Focal Cerebral Ischemia

BACKGROUND AND PURPOSE Magnetic resonance (MR) diffusion-weighted imaging (DWI), a noninvasive procedure, may play an important role in detecting and accurately localizing the extent of evolving infarction within the period immediately following stroke. We evaluated the sensitivity and specificity of DWI in detecting ischemia and compared a quantitative measure derived from the DWI, the apparent diffusion coefficient (ADC), with autoradiographic cerebral blood flow (CBF) in an experimental model of focal cerebral ischemia in rats. METHODS MR imaging data were obtained with a General Electric 4.7-T horizontal bore magnet CSI II system with self-shielded gradients. DWI was acquired within 41 +/- 6 minutes (mean +/- SD) after onset of ischemia and repeated at 169 +/- 14 minutes, followed by CBF determination at 237 +/- 21 minutes. DWI, ADC, and CBF images from each animal were then compared. RESULTS The sensitivities for detecting an abnormality at 1 and 3 hours for DWI were significantly different, and the sensitivity of 3-hour DWI did not differ from the CBF sensitivity of 99%. A mean +/- SD ADC threshold of 460 +/- 95 microns 2/s was defined as 45% higher than the low ADC in the ischemic core compared with the contralateral ADC. Subthreshold ADC area and ischemic area were significantly correlated (r2 = .69, P < .05). In 19 of 48 regions of interest classified as ischemic (< 35 mL.100 g-1.min-1) from both the 3-hour ADC and CBF images, 3-hour ADC correlated significantly with CBF (r2 = .27, n = 19, P < .05), whereas in the nonischemic regions ADC was inversely correlated with CBF. Several ischemic regions showed a sharp drop in ADC to 37% (P < .001, n = 5) compared with all other regions (n = 43) from 1 to 3 hours. CONCLUSIONS Because of the change in the sensitivity of detecting ischemia with DWI, the difference in correlation of CBF with ADC between ischemic and nonischemic cortex, and the presence of several regions in which ADC dropped to 37% from 1 to 3 hours, our data suggest that ADC values potentially can be used to monitor evolving infarction.

Pretransplant assessment of renal viability by phosphorus-31 magnetic resonance spectroscopy. Clinical experience in 40 recipient patients.

A group of 40 cadaveric kidneys was studied just prior to planned transplantation to further assess the applicability of 31P-MRS in the analysis of clinical renal transplant viability. Renal intracellular high-energy phosphorus metabolites (ATP [or NADP], phosphomonoester [PME] and inorganic phosphate [Pi]) and pH were measured noninvasively with MRS surface coils external to cold storage containers. Pretransplant MRS parameters were correlated with subsequent renal function in recipient patients (measured one week postoperatively by the need of dialysis, drop in serum creatinine, urine output, and 123I or 131I Hippuran assessed renal tubular function). ATP and NADP was detected in eleven kidneys and was significantly (P<0.001) associated with the best renal function posttransplantation. These kidneys also had the highest PME/Pi ratios (1.66–0.54), while lower ratios (0.36–0.10) were associated with prolonged acute tubular necrosis. The PME/Pi ratios significantly (P<0.0001) correlated with subsequent clinical renal function, whereas cold storage times (37±10 hr) or intracellular renal pH (6.53–7.91) did not. These preliminary data suggest that MRS is a noninvasive, nondestructive and sterile method for assessing clinical viability during hypothermic storage of human cadaver kidneys and the subsequent recovery of renal function postrenal transplantation.