Journal of Neuro-Oncology 14: 207-211, 1992. © 1992 Kluwer Academic Publishers. Printed in the Netherlands.
Laboratory Investigation
MR imaging of experimental meningeal melanomatosis in nude rats Jfinos Martos, 1,2 Dirk Petersen, 1 Uwe Klose, 1 Hermann Requardt, 3 Rainer Buchholz, 4 Petra Ohneseit, 5 Martin Schabet 6 and Karsten Voigt 1
1Department of Neuroradiology and 4Neurosurgery and 5Radiotherapy and °Neurology, University of Tiibingen, Hoppe Seyler Str. 3., D-7400 Tiibingen, Germany 2National Institute of Neurosurgery, Amerikai tit 57 H-1145 Budapest, Hungary 3Siemens Medical Systems, Henkestrasse 127, D-8520 Erlangen, Germany
Key words: leptomeningeal metastasis, B16 melanoma, magnetic resonance imaging, animal model Abstract
MR imaging of the rat brain has become an increasingly frequently used method in experimental neuroradiology. On a generally available 1.5 T whole body tomograph, supplemented with an individually made small coil and a special SE sequence we obtained fairly fine images of the structures of the rat brain. With gadolinium-DTPA, we were able to visualize posterior fossa and cervical leptomeningeal growth of intrathecally injected B16 melanoma in nude rats. Using MRI to follow experimental leptomeningeal metastasis, may provide a new means for diagnostic evaluation and preclinical testing of treatment modalities.
Introduction
Materials and methods
Magnetic resonance imaging (MRI) can improve the quality of animal experiments, and also help to reveal the histological background of MR findings. Rat brain imaging using a conventional whole body MR unit, provides suitable soft tissue contrast to distinguish the main structures of the brain and to visualize experimentally induced tumors or traumatic lesions [1, 2]. Leptomeningeal metastasis occurs in 5-30% of patients with breast or lung cancer, malignant melanoma, Non-Hodgkin's lymphoma, leukemia and primary malignant brain tumors. The prognosis of this complication is poor with a median survival of 2-7 month [3, 4]. Leptomeningeal metastasis of mouse B 16 melanoma in nude rats has been established as a new animal model for preclinical testing of several route of therapy [5]. MRI is shown to be a suitable means to follow up in vivo tumor growth.
Six female Rowett nude rats (100-120mg) were obtained from the Zentrale Tierzuchtanstalt in Hannover and housed under normal non-sterile conditions. The rats were anesthetized with 62.5 mg/kg ketanest (Ketavet TM,Parke Davis, Berlin, FRG) and 2.87mg/kg xylazin (Rompun TM, Bayer, Leverkusen, FRG) and intracisternally injected with 10 4 m o u s e B16 melanoma cells as previously described [5]. Animals were investigated with MRI when neurological signs first developed. Immediately after imaging, rats were sacrificed with an overdose of ketanest and xylazine for macroscopic and histologic evaluation. For the MR investigation the rats were anesthetized with a simple Halothane/N20/O2 open system. Imaging was performed with a 7.5 cm internal diameter, 13 cm long resonator, and used in a 1.5 T whole-body imager (Magnetom a, Siemens, Erlangen, FRG). To achieve 6 cm field of view on the
208
Fig. 1. Ventral (a) and dorsal (b) aspect of brain and spinal cord showing mass growth of B16 melanoma from the cisterna magna to the midcervical region and patchy growth along the brain stem and spinal cord.
256 x 256 image matrix, a spin-echo sequence with a read-out time of 11.5 msec was chosen, because the imager was equipped with maximum of 10 mT/m gradient coils. Parameters of the SE sequence usually included 3 mm slice thickness with an interslice skip of 1.5mm, TR/TE was 500/ 28 msec for the Tl-weighted images. As a contrast material, 0.5 mmol/kg Gd-DTPA (Magnevist R, Schering, Berlin, FRG) was prepared in a concentration of 0.25 mmol/ml in normal saline and injected via the tail vein. The macroscopic and microscopic sections of the brain and spinal cord were cut parallelly to the MR images and documented photographically. Histological evaluation was done on slides stained with haematoxylin and eosin after embedding in paraffin.
Fig. 2. Infiltration of the cerebellar cortex with tumor cells (H.E. × 130).
Results
Clinical and neuropathological data are detailed in Table 1. Figure 1-4 are from rat 2. All six animals
Fig. 3. Sagittal Tl-weighted (SE 500/28) precontrast image shows space occupying lesion in the cisterna magna (a). Sagittal (b) and axial (c) Gd-DTPA enhanced images demonstrate better the bulky tumor mass and reveal a small enhanced region at the ventral margin of the cervical spinal cord (arrow) corresponding to patchy tumor growth (Fig. 1).
209
Fig. 4. Gd contrast enhanced coronal Tl-weighted (SE 500/28) serial images show the enlarged ventricles (a) and the tumor in the cisterna magna as well as its great mass effect on the brain stern (c). The macroscopic slices, which are corresponding to the coronal MR images, show the enlarged ventricles (b) and the black tumor mass compressing the pons and medulla oblongata (d).
exhibited marked tumor growth in the cisterna magna. Leptomeningeal growth was less intense or absent in the basal frontal cisterns and along the spinal cord (Fig. 1). At sites of mass or multilayer growth, the adjoining cerebellar cortex or super-
ficial parenchyma of the brain stem and spinal cord were severely infiltrated (Fig. 2). The tumor mass in the cisterna magna was always clearly visible on precontrast Tl-weighted images as a space-occupying isointense lesion, but it
Table 1. Clinical, MR and neuropathological findings of nude rats inoculated with 104 B16 melanoma cells Rat
Clinical findings
M R I findings
Symptoms Survival time (days)
Hydrocephalus
Neuropathological findings
no.
1 2 3 4 5 6
TP TP PP TP TP TP
20 22 18 17 18 21
+++ +++ +++ ++ + -
Tumor volume (mm 3)
323.97 245.43 141.11 116.63 78.34 77.15
Tumor growth Cisterna magna Basal cisternae
Spinal upper
Spinal lower
+++ +++ +++ +++ +++ +++
+ + + + +
+ + -
+ + + + -
Symptoms: TP = tetraparesis, PP = paraparesis of hindlegs; Survival time = free of symptoms; Hydrocephalus: + + + = extensive, + + = moderate, + = beginning, - = absent; T u m o r volume: calculated from an ovoid in the cisterna magna; Tumor growth: + + + = mass growth, + = unilayer/spotty multilayer, - = no growth.
210 displayed intense enhancement and was better defined after Gd-DTPA administration (Fig. 3, 4a). On the postcontrast images some small enhanced spots were also seen along the cervical spinal cord (Fig. 3b). The patchy growth in the basal cisterns and the lower spinal leptomeningeal spread was not observed on the MR images. The mass effect upon the fourth ventricle and the histologically documented infiltration of the CSF pathways caused hydrocephalus internus in five rats. The enlarged aqueduct and supratentorial ventricles were dearly depicted on the MR images (Fig. 4a, b).
Discussion In patients with leptomeningeal metastasis, the Gd-DTPA enhanced MRI is a worthwhile method to support diagnosis and to follow up disease. It is more sensitive than contrast enhanced CT of the brain and equally sensitive as myelography in detection of spinal seeding [7-9]. Smaller lesions, however, may be inapparent and the distinction may be difficult between enhancing tumor lesions and enhancement due to non-specific blood-brain and blood-CSF barrier disturbances. Any process that effects meningeal irritation can also cause visible meningeal thickening [10, 11]. Therefore experimental models are required to clarify the histopathological correlations and to allow experimental follow-up studies. Frank et al. [12] were the first to use MRI in an experimentally induced meningeal carcinomatosis with VX2 rabbit carcinoma cells on an 0.5T MR unit. They found enhancement of tumor plaques as seen in patients, but they also stressed the problem of plaque size and non-specific leptomeningeal enhancement. High field strength whole body MR units, supplemented with a suitable small coil also make rat brain imaging possible. Runge et al. reported about rat brain MRI following stereotactic implantation of C6 glioma (1) and 9 L gliosarcoma cells (2) in Fisher rats on a 1T MR imager. After injection of 9 L cells into the basal ganglia, meningeal seeding was detected on Gd-DTPA enhanced images sur-
rounding the cerebrum and cerebellum. MRI of meningeal melanomatosis induced in nude rats is first reported here. The well-characterized, fast-growing B16 melanoma is advantageous for MRI, because it provides well detectable mass or multilayer leptomeningeal growth at sites of inoculation and adjoining subarachnoid space. In our nude rat model with B16 melanoma cells [5, 6] we found a good imagehistopathological correlation without non-specific enhancement. Gd DTPA enhanced MRI clearly defined space-occupying lesion in the cisterna magna and small lesions along the cervical spinal cord. The patchy growth in the basal cisterns and the lower spinal leptomeningeal spread was not observed on the MR images due to the small size of these lesions and the respiratory movement artifacts on the spinal scans. With this in minde, MRI may still be a suitable means in monitoring disease when studying treatment modalities in this model.
Acknowledgment This study was supported by Schering Comp. AG, Berlin.
References 1. Runge VM, Jacobson S, Wood ML, Kaufman D, Adelman LS: MR imaging of rat brain glioma: Gd-DTPA versus Gd-DOTA. Radiology 166: 835-838, 1988 2. Runge VM, Gelblum DY, Jacobson S: Gd HP-DO3A Experimental evaluation in brain and renal MR. Magnetic Resonance Imaging 9: 79-87, 1991 3. Bleyer WA, Byrne TN: Leptomeningeal cancer in leukemia and solid tumors. Curr Probl Cancer 12: 181-238, 1988 4. Schabet M, Bamberg M, Dichgans J: Diagnose und therapie der meningosis neoplastica. Nervenarzt 1992 (in press) 5. Schabet M, Wieth01ter H, Meier D, Birchmeier W: Experimental meningeal melanomatosis. Strahlenther Onkol 165 (7): 491--492, 1989 6. Schabet M, Ohneseit P, Buchholz R, Santo-H01tje L, Schmidberger H: Intrathecal ACNU treatment of B16 melanoma leptomeningeal metastasis in a new athymic rat model. J Neuro-Oncol 1992 (in press) 7. Davis PC, Friedman NC, Fry SM, Malko JA, Hoffman JC, JR., Braun IF: Leptomeningeal metastasis: MR imaging. Radiology 163: 449-454, 1987
211 8. Tyrell RL, Bundschuh CV, Modic MT: Dural carcinomatosis: MR demonstration. J Comput Assist Tomogr 11(2): 329-332, 1987 9. Sze G, Abramson A, Krol G, Liu D, Amster J, Zimmerman RD, Deck MD: Gadolinium-DTPAin the evaluation of intradural extramedullary spinal disease. AJR 150: 911921, 1988 10. Sze G, Soletsky S, Bronen R, Krol G: MR imaging of the cranial meninges with emphasis on contrast enhancement and meningeal carcinomatosis. AJNR 10: 965-975, 1989 11. Rodesch G, Van Bogaert P, Mavroudakis N, Parizel PM, Martin J-J, Segebarth C, Van Vyve M, Baleriaux D, Hildebrand J: Neuroradiologic findings in leptomeningeal carci-
nomatosis: the value interest of gadolinium-enhanced MRI. Neuroradiology 32: 26--32, 1990 12. Frank JA, Girton M, Dwyer AJ, Wright DC, Cohen PJ, Doppman JL: Meningeal carcinomatosis in the VX2 rabbit tumor model: detection with Gd-DTPA enhanced MR imaging. Radiology 167: 825-829, 1988
Address for offprints: J. Martos, Department of Neuroradiology, University of Tiibingen, Hoppe Seyler Str. 3, D-7400 Ttibingen, Germany
Laboratory Investigation
MR imaging of experimental meningeal melanomatosis in nude rats Jfinos Martos, 1,2 Dirk Petersen, 1 Uwe Klose, 1 Hermann Requardt, 3 Rainer Buchholz, 4 Petra Ohneseit, 5 Martin Schabet 6 and Karsten Voigt 1
1Department of Neuroradiology and 4Neurosurgery and 5Radiotherapy and °Neurology, University of Tiibingen, Hoppe Seyler Str. 3., D-7400 Tiibingen, Germany 2National Institute of Neurosurgery, Amerikai tit 57 H-1145 Budapest, Hungary 3Siemens Medical Systems, Henkestrasse 127, D-8520 Erlangen, Germany
Key words: leptomeningeal metastasis, B16 melanoma, magnetic resonance imaging, animal model Abstract
MR imaging of the rat brain has become an increasingly frequently used method in experimental neuroradiology. On a generally available 1.5 T whole body tomograph, supplemented with an individually made small coil and a special SE sequence we obtained fairly fine images of the structures of the rat brain. With gadolinium-DTPA, we were able to visualize posterior fossa and cervical leptomeningeal growth of intrathecally injected B16 melanoma in nude rats. Using MRI to follow experimental leptomeningeal metastasis, may provide a new means for diagnostic evaluation and preclinical testing of treatment modalities.
Introduction
Materials and methods
Magnetic resonance imaging (MRI) can improve the quality of animal experiments, and also help to reveal the histological background of MR findings. Rat brain imaging using a conventional whole body MR unit, provides suitable soft tissue contrast to distinguish the main structures of the brain and to visualize experimentally induced tumors or traumatic lesions [1, 2]. Leptomeningeal metastasis occurs in 5-30% of patients with breast or lung cancer, malignant melanoma, Non-Hodgkin's lymphoma, leukemia and primary malignant brain tumors. The prognosis of this complication is poor with a median survival of 2-7 month [3, 4]. Leptomeningeal metastasis of mouse B 16 melanoma in nude rats has been established as a new animal model for preclinical testing of several route of therapy [5]. MRI is shown to be a suitable means to follow up in vivo tumor growth.
Six female Rowett nude rats (100-120mg) were obtained from the Zentrale Tierzuchtanstalt in Hannover and housed under normal non-sterile conditions. The rats were anesthetized with 62.5 mg/kg ketanest (Ketavet TM,Parke Davis, Berlin, FRG) and 2.87mg/kg xylazin (Rompun TM, Bayer, Leverkusen, FRG) and intracisternally injected with 10 4 m o u s e B16 melanoma cells as previously described [5]. Animals were investigated with MRI when neurological signs first developed. Immediately after imaging, rats were sacrificed with an overdose of ketanest and xylazine for macroscopic and histologic evaluation. For the MR investigation the rats were anesthetized with a simple Halothane/N20/O2 open system. Imaging was performed with a 7.5 cm internal diameter, 13 cm long resonator, and used in a 1.5 T whole-body imager (Magnetom a, Siemens, Erlangen, FRG). To achieve 6 cm field of view on the
208
Fig. 1. Ventral (a) and dorsal (b) aspect of brain and spinal cord showing mass growth of B16 melanoma from the cisterna magna to the midcervical region and patchy growth along the brain stem and spinal cord.
256 x 256 image matrix, a spin-echo sequence with a read-out time of 11.5 msec was chosen, because the imager was equipped with maximum of 10 mT/m gradient coils. Parameters of the SE sequence usually included 3 mm slice thickness with an interslice skip of 1.5mm, TR/TE was 500/ 28 msec for the Tl-weighted images. As a contrast material, 0.5 mmol/kg Gd-DTPA (Magnevist R, Schering, Berlin, FRG) was prepared in a concentration of 0.25 mmol/ml in normal saline and injected via the tail vein. The macroscopic and microscopic sections of the brain and spinal cord were cut parallelly to the MR images and documented photographically. Histological evaluation was done on slides stained with haematoxylin and eosin after embedding in paraffin.
Fig. 2. Infiltration of the cerebellar cortex with tumor cells (H.E. × 130).
Results
Clinical and neuropathological data are detailed in Table 1. Figure 1-4 are from rat 2. All six animals
Fig. 3. Sagittal Tl-weighted (SE 500/28) precontrast image shows space occupying lesion in the cisterna magna (a). Sagittal (b) and axial (c) Gd-DTPA enhanced images demonstrate better the bulky tumor mass and reveal a small enhanced region at the ventral margin of the cervical spinal cord (arrow) corresponding to patchy tumor growth (Fig. 1).
209
Fig. 4. Gd contrast enhanced coronal Tl-weighted (SE 500/28) serial images show the enlarged ventricles (a) and the tumor in the cisterna magna as well as its great mass effect on the brain stern (c). The macroscopic slices, which are corresponding to the coronal MR images, show the enlarged ventricles (b) and the black tumor mass compressing the pons and medulla oblongata (d).
exhibited marked tumor growth in the cisterna magna. Leptomeningeal growth was less intense or absent in the basal frontal cisterns and along the spinal cord (Fig. 1). At sites of mass or multilayer growth, the adjoining cerebellar cortex or super-
ficial parenchyma of the brain stem and spinal cord were severely infiltrated (Fig. 2). The tumor mass in the cisterna magna was always clearly visible on precontrast Tl-weighted images as a space-occupying isointense lesion, but it
Table 1. Clinical, MR and neuropathological findings of nude rats inoculated with 104 B16 melanoma cells Rat
Clinical findings
M R I findings
Symptoms Survival time (days)
Hydrocephalus
Neuropathological findings
no.
1 2 3 4 5 6
TP TP PP TP TP TP
20 22 18 17 18 21
+++ +++ +++ ++ + -
Tumor volume (mm 3)
323.97 245.43 141.11 116.63 78.34 77.15
Tumor growth Cisterna magna Basal cisternae
Spinal upper
Spinal lower
+++ +++ +++ +++ +++ +++
+ + + + +
+ + -
+ + + + -
Symptoms: TP = tetraparesis, PP = paraparesis of hindlegs; Survival time = free of symptoms; Hydrocephalus: + + + = extensive, + + = moderate, + = beginning, - = absent; T u m o r volume: calculated from an ovoid in the cisterna magna; Tumor growth: + + + = mass growth, + = unilayer/spotty multilayer, - = no growth.
210 displayed intense enhancement and was better defined after Gd-DTPA administration (Fig. 3, 4a). On the postcontrast images some small enhanced spots were also seen along the cervical spinal cord (Fig. 3b). The patchy growth in the basal cisterns and the lower spinal leptomeningeal spread was not observed on the MR images. The mass effect upon the fourth ventricle and the histologically documented infiltration of the CSF pathways caused hydrocephalus internus in five rats. The enlarged aqueduct and supratentorial ventricles were dearly depicted on the MR images (Fig. 4a, b).
Discussion In patients with leptomeningeal metastasis, the Gd-DTPA enhanced MRI is a worthwhile method to support diagnosis and to follow up disease. It is more sensitive than contrast enhanced CT of the brain and equally sensitive as myelography in detection of spinal seeding [7-9]. Smaller lesions, however, may be inapparent and the distinction may be difficult between enhancing tumor lesions and enhancement due to non-specific blood-brain and blood-CSF barrier disturbances. Any process that effects meningeal irritation can also cause visible meningeal thickening [10, 11]. Therefore experimental models are required to clarify the histopathological correlations and to allow experimental follow-up studies. Frank et al. [12] were the first to use MRI in an experimentally induced meningeal carcinomatosis with VX2 rabbit carcinoma cells on an 0.5T MR unit. They found enhancement of tumor plaques as seen in patients, but they also stressed the problem of plaque size and non-specific leptomeningeal enhancement. High field strength whole body MR units, supplemented with a suitable small coil also make rat brain imaging possible. Runge et al. reported about rat brain MRI following stereotactic implantation of C6 glioma (1) and 9 L gliosarcoma cells (2) in Fisher rats on a 1T MR imager. After injection of 9 L cells into the basal ganglia, meningeal seeding was detected on Gd-DTPA enhanced images sur-
rounding the cerebrum and cerebellum. MRI of meningeal melanomatosis induced in nude rats is first reported here. The well-characterized, fast-growing B16 melanoma is advantageous for MRI, because it provides well detectable mass or multilayer leptomeningeal growth at sites of inoculation and adjoining subarachnoid space. In our nude rat model with B16 melanoma cells [5, 6] we found a good imagehistopathological correlation without non-specific enhancement. Gd DTPA enhanced MRI clearly defined space-occupying lesion in the cisterna magna and small lesions along the cervical spinal cord. The patchy growth in the basal cisterns and the lower spinal leptomeningeal spread was not observed on the MR images due to the small size of these lesions and the respiratory movement artifacts on the spinal scans. With this in minde, MRI may still be a suitable means in monitoring disease when studying treatment modalities in this model.
Acknowledgment This study was supported by Schering Comp. AG, Berlin.
References 1. Runge VM, Jacobson S, Wood ML, Kaufman D, Adelman LS: MR imaging of rat brain glioma: Gd-DTPA versus Gd-DOTA. Radiology 166: 835-838, 1988 2. Runge VM, Gelblum DY, Jacobson S: Gd HP-DO3A Experimental evaluation in brain and renal MR. Magnetic Resonance Imaging 9: 79-87, 1991 3. Bleyer WA, Byrne TN: Leptomeningeal cancer in leukemia and solid tumors. Curr Probl Cancer 12: 181-238, 1988 4. Schabet M, Bamberg M, Dichgans J: Diagnose und therapie der meningosis neoplastica. Nervenarzt 1992 (in press) 5. Schabet M, Wieth01ter H, Meier D, Birchmeier W: Experimental meningeal melanomatosis. Strahlenther Onkol 165 (7): 491--492, 1989 6. Schabet M, Ohneseit P, Buchholz R, Santo-H01tje L, Schmidberger H: Intrathecal ACNU treatment of B16 melanoma leptomeningeal metastasis in a new athymic rat model. J Neuro-Oncol 1992 (in press) 7. Davis PC, Friedman NC, Fry SM, Malko JA, Hoffman JC, JR., Braun IF: Leptomeningeal metastasis: MR imaging. Radiology 163: 449-454, 1987
211 8. Tyrell RL, Bundschuh CV, Modic MT: Dural carcinomatosis: MR demonstration. J Comput Assist Tomogr 11(2): 329-332, 1987 9. Sze G, Abramson A, Krol G, Liu D, Amster J, Zimmerman RD, Deck MD: Gadolinium-DTPAin the evaluation of intradural extramedullary spinal disease. AJR 150: 911921, 1988 10. Sze G, Soletsky S, Bronen R, Krol G: MR imaging of the cranial meninges with emphasis on contrast enhancement and meningeal carcinomatosis. AJNR 10: 965-975, 1989 11. Rodesch G, Van Bogaert P, Mavroudakis N, Parizel PM, Martin J-J, Segebarth C, Van Vyve M, Baleriaux D, Hildebrand J: Neuroradiologic findings in leptomeningeal carci-
nomatosis: the value interest of gadolinium-enhanced MRI. Neuroradiology 32: 26--32, 1990 12. Frank JA, Girton M, Dwyer AJ, Wright DC, Cohen PJ, Doppman JL: Meningeal carcinomatosis in the VX2 rabbit tumor model: detection with Gd-DTPA enhanced MR imaging. Radiology 167: 825-829, 1988
Address for offprints: J. Martos, Department of Neuroradiology, University of Tiibingen, Hoppe Seyler Str. 3, D-7400 Ttibingen, Germany
