Animal model of metastatic pheochromocytoma: evaluation by MRI and PET.

OBJECTIVE: The development of metastatic pheochromocytoma animal model provides a unique opportunity to study the physiology of these rare tumors and to evaluate experimental treatments. Here, we describe the use of small animal imaging techniques to detect, localize and characterize metastatic lesions in nude mice. METHODS: Small animal positron emission tomography (PET) imaging and magnetic resonance imaging (MRI) were used to detect metastatic lesions in nude mice following intravenous injection of mouse pheochromocytoma cells. [18F]-6-fluoro-dopamine ([18F]-DA) and [18F]-L-6-fluoro-3,4-dihydroxyphenylalanine, which are commonly used for localization of pheochromocytoma lesions in clinical practice, were selected as radiotracers to monitor metastatic lesions by PET. RESULTS: MRI was able to detect liver lesions as small as 0.5mm in diameter. Small animal PET imaging using [18F]-DA and [18F]-DOPA detected liver, adrenal gland, and ovarian lesions. CONCLUSION: We conclude that MRI is a valuable technique for tumor growth monitoring from very early to late stages of tumor progression and that animal PET confirmed localization of metastatic pheochromocytoma in liver with both radiotracers.

Abdominal MR imaging with a volumetric interpolated breath-hold examination.

PURPOSE: To compare a T1-weighted, three-dimensional (3D), gradient-echo (GRE) sequence for magnetic resonance (MR) imaging of the body (volumetric interpolated breath-hold examination, or VIBE) with a two-dimensional (2D) GRE breath-hold equivalent. MATERIALS AND METHODS: Twenty consecutive patients underwent 1.5-T MR imaging. The examinations included pre- and postcontrast (20 mL gadopentetate dimeglumine) fat-saturated 2D GRE breath-hold imaging and fat-saturated volumetric interpolated breath-hold imaging before, during (arterial phase), and after injection, with thin (2-mm source images) and thick (8-mm reconstruction images) sections. The three images were compared qualitatively and quantitatively (signal-to-noise ratio [SNR] and contrast-to-noise ratio [CNR]). RESULTS: Qualitatively, the 2-mm source images had poorer pancreatic edge definition on precontrast images compared with the other two data sets (P < .05). On gadolinium-enhanced images, scores for clarity of pancreatic edge, number of vessels visualized, and arterial ghosting were significantly lower for the postcontrast 2D GRE images. Quantitatively, SNR measurements in the liver, aorta, and renal cortex on pre- and postcontrast images were significantly higher for the 8-mm reconstruction images than for the 2D GRE or 2-mm source images (P < .05). Aorta-to-fat CNR was significantly higher on the 8-mm reconstruction images. CONCLUSION: Fat-saturated volumetric interpolated breath-hold images have quality comparable to that of conventional fat-saturated 2D GRE images.

T2-weighted MRI of the uterus: fast spin echo vs. breath-hold fast spin echo.

This study compared one routine T2-weighted fast spin echo (T2FSE) sequence with a breath-hold T2FSE (BH T2FSE) sequence of the female pelvis for image quality, uterine anatomy, lesion detection, and signal intensity measurements. Thirty-two consecutive women (mean age 41.7 years) were imaged at 1.5 T with one high-resolution routine T2FSE sequence and one BH T2FSE sequence in the sagittal plane as part of comprehensive pelvic magnetic resonance imaging. The different image sets were rated separately for imaging characteristics (overall image quality, uterine anatomy definition, lesion detection, and free fluid conspicuity) and then compared side by side. The image sets were also compared for artifacts (ghosting, blurring, pulsatility, and chemical shift misregistration). Signal-to-noise (S/N) and signal difference-to-noise (SD/N) ratios were calculated for the different uterine zones, uterine abnormalities, free fluid, rectus abdominis muscle, and bladder. Contrast-to-noise ratios (CNRs) were calculated for uterine abnormalities. Twenty-eight uterine abnormalities were detected in 20 patients and included leiomyomata (13 patients), adenomyosis (7 patients), benign endometrial polyps (6 patients), endometrial carcinoma (1 patient), and pregnancy (1 patient). BH T2FSE was superior or equivalent to T2FSE for overall image quality in 23/32 patients (71.8%), uterine anatomy definition in 19/32 patients (59.3%), and lesion detection in 13/20 patients (65%). BH T2FSE performed less well than T2FSE for free fluid conspicuity in 5/5 (100%) patients. BH T2FSE was equivalent to or less affected than T2FSE for ghosting artifact in 24/32 patients (75%) and blurring artifact in 29/32 patients (90.6%). Pulsatility and chemical shift artifacts were not problematic for either image set. S/N and SD/N were higher for all BH T2FSE determinations compared with T2FSE. For the endometrium, junctional zone, myometrium, and bladder, these differences were statistically significant. There were no statistically significant differences for CNR between the two image sets, although BH T2FSE values for leiomyomata, adenomyosis, and abnormal endometria were higher than those calculated for T2FSE. All pathology detected with T2FSE was detected on BH T2FSE despite the breath-hold sequence's inherently poorer spatial resolution compared with the non-breath-hold sequence. BH T2FSE may be able to replace T2FSE for some uterine applications with a substantial time savings.

Quantitation of the susceptibility difference between trabecular bone and bone marrow: experimental studies.

In this study we quantify the effects of different relaxation mechanisms on the signal intensity in gradient-echo images of tissue such as bone marrow in the presence of trabecular bone. The susceptibility difference between trabecular bone and soft tissue produces distortions in the magnetic lines of force which induce strong inhomogeneities in the static magnetic field. Diffusion of tissue protons in such magnetic field gradients produce a shortening of the transverse relaxation time T2, while the dephasing of the transverse magnetization due to susceptibility differences produces a shortening of the apparent relaxation time T2* as demonstrated in gradient-echo images. We have used specimens of dried human vertebrae with different bone densities immersed in either saline to simulate tissue water or an emulsion of oil and water to simulate bone marrow to quantify these relaxation mechanisms in vitro. We have measured the MR relaxation times T1, T2, and T2* of protons within the trabecular spaces and correlated their variations with trabecular bone density. We have found that in vitro, at 1.5 T, the relaxation times T1 and T2 do not show significant variations with bone density and there are no significant contributions to the transverse relaxation rate due to the diffusion of tissue water in the magnetic field gradients. However, the relaxation rate, 1/T2*, of saline in the presence of trabecular bone increases at a rate of 0.2 s-1/mg/cc due to the dephasing of the transverse magnetization in the magnetic field inhomogeneities. Similar bone density-related T2* variations were observed for fat protons within the trabeculae where the chemical-shift-induced modulations of signal intensity in an oil-water emulsion have been separated from the susceptibility-induced relaxation effects. In addition, we have verified these effects in vivo and quantified in vivo variations in fat and water relaxation rates of bone marrow in the epiphysis and diaphysis in the appendicular skeleton of normal volunteers and found that both fat and water T2* are shorter in the epiphysis compared to the diaphysis, which correlates well with previous observations.

Multiple short-echo (2.5-ms) quantitation of in vivo sodium T2 relaxation.

The MR behavior of the sodium-23 nucleus in vivo is a complex problem which has generated considerable interest over the last 20 years. Early studies on excised tissue samples revealed that the sodium nucleus exhibited a two-component T2 relaxation. This biexponential T2 relaxation was characterized by a short component with a T2 = 0.7-4.8 ms, and a long component with a T2 = 7.0-26.0 ms. We have developed a 3D pulse sequence capable of performing multiple Hahn echo in vivo sodium-23 imaging at echo times as short as 2.5 ms. This sequence obtains the shorter spin echo times by presaturating the spins outside of the desired imaging region, allowing the use of nonselective rf pulses. Using this sequence we have been able to quantify the long and short T2 components of normal brain tissue, vitreous humor of the eye, and a rabbit VX-2 carcinoma. We found that gray matter and white matter of normal brain have a monoexponential T2 relaxation with T2 = 17.6 +/- 2.4 ms. The vitreous humor T2 relaxation is also monoexponential with T2 = 56.8 +/- 2.1 ms. However, we find that some of the rabbit VX2 carcinomas exhibit a biexponential T2 decay with a short component of 3.3 +/- 4.6 ms and a long component of 22.0 +/- 9.0 ms.

Characteristics of bradykinin and TPA increases in the PGE2 levels of human urothelial cells.

Prostaglandins play a potential key role in the pathogenesis of urinary bladder cancer. Bradykinin and TPA increases in prostaglandin (PG)E2 levels were compared in primary cultures of human urothelial cells. Increased PGE2 levels were dependent upon the dose of TPA and were not apparent until 30-60 min after addition of TPA, with larger increases occurring between 60 and 120 min. Stimulation was inhibited by cycloheximide. Addition of arachidonic acid to TPA-stimulated cells increased PGE2 to a level similar to that seen in arachidonic acid-stimulated controls, and this level was not altered by cycloheximide. In contrast to TPA, the bradykinin-increased PGE2 levels were maximal at 5 min (the earliest time-point assessed) and were not inhibited by cycloheximide. Increases in PGE2 levels by both TPA and bradykinin required calcium. Excessive stimulation by TPA resulted in a desensitization to subsequent stimulation by TPA, but not bradykinin. Combination of TPA with bradykinin produced at least an additive effect on PGE2 levels. Both agonists increased the release of [3H]arachidonic acid over a time-course similar to their PGE2 response. Bradykinin and TPA appear to increase PGE2 levels by enhancing arachidonic acid availability through separate phospholipase pathways. Thus, human urothelial cells exhibit similar, but yet distinct profiles for prostaglandin stimulation by TPA and bradykinin.

Cortical interstitial cell interactions induce sensitivity of hydronephrotic kidney to bradykinin.

The mechanism of the increased prostaglandin production and induction of sensitivity to bradykinin by the cortex of the hydronephrotic rabbit kidney was investigated using tissue culture techniques. Cortical interstitial cells from normal, unilaterally hydronephrotic and contralateral kidneys were grown in tissue culture. Cells derived from hydronephrotic kidneys, but not normal or contralateral, increased PGE2 production when incubated with bradykinin. Of the two cell types, fibroblasts and macrophages, grown from hydronephrotic explants, neither increased prostaglandin production when grown alone in tissue culture. Recombining the two cell types restored bradykinin responsiveness. Bradykinin responsiveness could be induced in either normal or contralateral cell cultures when macrophages from the hydronephrotic kidney were added to cultures of cells from normal or contralateral cortex. The data indicate unique characteristics of hydronephrotic macrophages are involved in the induction of bradykinin responsiveness in the cortex of the ureter-ligated kidney.

Eicosanoid synthesis by cultured human urothelial cells: potential role in bladder cancer.

Prostaglandin (PG) H synthase and eicosanoid products of arachidonic acid metabolism have been implicated in several steps in the carcinogenic process. This study assessed these parameters using primary cultures of human urothelial cells. To determine the possible presence of permeability barriers to agonist stimulation, incubations were performed with adherent cells in the presence or absence of thioglycolate pretreatment or with cell suspensions. No evidence for permeability barriers was observed. With adherent cells in the absence of thioglycolate, radioimmunoassayable PGE2 was stimulated by epinephrine less than 12-O-tetradecanoylphorbol-13-acetate = thrombin less than bradykinin = A23187 much less than arachidonic acid. Tumor promoters but not non-tumor promoters stimulated PGE2 synthesis. 1-Oleoyl-2-acetylglycerol which like 12-O-tetradecanoylphorbol-13-acetate activates protein kinase C also increased PGE2 synthesis. Cells prelabeled with [14C]arachidonic acid were exposed to agonists and the profile of eicosanoids synthesized was assessed by high performance liquid chromatography. With bradykinin, the pattern of eicosanoids synthesized was 6-keto-PGE1 alpha (12% of total 14C label), thromboxane B2 (0.4%), PGF2 alpha (1.7%), PGE2 (18%), PGD2 (1%), leukotrienes (0.4 to 1%), 12-hydroxy-5,8,10-heptadecatrienoic acid (3%), 15-hydroxy-5,8,11,13-eicosatetraenoic acid (4%), 12-hydroxy-5,8,10,14-eicosatetraenoic acid (0%) and 5-hydroxy-5,8,12,14-eicosatetraenoic acid (2%). Thus, human urothelial cells contain both prostaglandin H synthase and lipoxygenase pathways with the former being more prominent. These pathways may participate in urinary bladder carcinogenesis.

Effect of phosphodiesterase inhibitors on bradykinin-mediated prostaglandin E2 and cyclic AMP synthesis in renal papillary collecting tubule cells.

Isolated rabbit renal papillary collecting tubule cells were used to examine the effects of phosphodiesterase inhibitors on intracellular cyclic AMP and prostaglandin synthesis. Experiments performed on confluent primary tissue cultures demonstrated that bradykinin increases intracellular cyclic AMP by a prostaglandin-dependent mechanism. Phosphodiesterase inhibitors induced a dose-dependent decrease in bradykinin-stimulated prostaglandin synthesis. Fifty percent inhibition occurred with approximately 0.7 mM 3-isobutyl-1-methylxanthine (IBMX). Inhibition was found to be reversible. IBMX did not inhibit bradykinin-induced prostaglandin synthesis as a result of increased intracellular cyclic AMP. The nonmethylxanthine phosphodiesterase inhibitor RO 20-1724 also reduced bradykinin-stimulated prostaglandin synthesis. IBMX inhibited calcium-ionophore-A23187-induced prostaglandin synthesis but did not inhibit arachidonic acid stimulation of prostaglandin synthesis. The data demonstrate that bradykinin increased renal papillary collecting tubule cell cyclic AMP in a prostaglandin-dependent manner. Based on the data presented, phosphodiesterase inhibitors act to decrease arachidonic acid availability for prostaglandin synthesis, independent of changes in cellular cyclic AMP content.

Effects of Mycobacterium bovis (strain BCG) on the interstitial cells of hydronephrotic, contralateral, and normal rabbit kidneys.

Studies were undertaken to determine the effect of viable organisms of Mycobacterium bovis, strain Bacillus Calmette Guerin (BCG), on cell growth characteristics and phagocytic properties of cells from surgically-induced unilaterally hydronephrotic, contralateral, and normal rabbit kidneys. A single intravenous administration of 8 X 10(8) BCG organisms was given at the time of ureteral ligation. Four days after injection, explants were removed from the hydronephrotic, contralateral, and normal kidneys. Two cell types, fibroblasts and mononuclear phagocytes, grew from these explants. BCG caused a marked increase in the rate of growth of cells from the hydronephrotic and contralateral kidneys. There was no measurable effect of BCG on cells from the normal kidney.

Renal disease profoundly alters cortical interstitial cell function.

Interstitial cells were cultured from explants of the unilaterally hydronephrotic, contralateral, and normal kidneys. Two types of cells were identified in culture, macrophages, and cells which were tentatively identified as fibroblasts. Cells grew at a significantly faster rate in hydronephrotic compared to contralateral or normal kidneys. Cells from the hydronephrotic kidney increased prostaglandin (PG)E2 production in response to bradykinin. Cells from contralateral and normal renal cortex did not increase PGE2 production in response to bradykinin. These results indicate hydronephrosis induces functional changes in interstitial cells cultured from the cortex of hydronephrotic compared to contralateral and normal kidneys. The induction of increased PGE2 synthesis and bradykinin responsiveness in hydronephrotic cortex could be related to the exaggerated prostaglandin synthesis known to occur in hydronephrotic cortex. In hydronephrosis, cortical interstitial cells elaborate increased amounts of substances such as prostaglandins which have the capacity to modulate important parameters of renal function.

T-cell hybridoma production of macrophage activation factor (MAF) I. Separation of MAF from interferon gamma.

A T cell hybridoma was developed that produced macrophage activation factor (MAF) but no gamma interferon (IFN gamma). Hybridoma MAF was produced after stimulation with either concanavalin A (Con A) or phytohemagglutinin (PHA). Dose-response experiments showed that 1.5 microgram/ml Con A provided maximum MAF production with equivalent levels of MAF produced at Con A concentrations up to 6 microgram/ml. Time-course studies showed that maximum MAF activity was observed 48 hours after mitogen stimulation, and titration of MAF-containing supernatants showed that maximal MAF activity was observed at 1:4 dilutions with significant activity present at dilutions as great as 1:64. No IFN gamma activity was detectable at any supernatant dilution.

The activation of cloned macrophages by concanavalin A for tumoricidal effect: assessment of tumor cell cytotoxicity by a clonogenic assay.

The activation of pure populations of cloned peritoneal macrophages for tumor cell cytotoxicity by Con A was demonstrated using a recently developed tumor cell clonogenic assay. Cloned macrophages were rendered cytotoxic by Con A at concentrations above 20 micrograms/ml. The tumor cell cytotoxicity was caused mainly by the tumoricidal activity of the Con A-activated cloned macrophages. Increasing The Con A-activation time from 24 hours to 48 hours and 72 hours heightened the cytotoxic activity of cloned macrophages. Cloned macrophages incubated with con A for only 2 hours possessed no cytotoxic effect. Culture fluid from cultures of activated macrophages exerted no tumor cell cytotoxicity. Alpha-methyl-D-mannoside, a specific receptor-binding inhibitor for Con A, was capable of blocking the activation of macrophages for cytotoxicity at 0.01 M concentration.