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DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF RAT TESTES: A METHOD FOR EARLY DETECTION OF ISCHEMIA
0022-5347/01/1666-2542/0 THE JOURNAL OF UROLOGY® Copyright © 2001 by AMERICAN UROLOGICAL ASSOCIATION, INC.®
Vol. 166, 2542–2544, December 2001 Printed in U.S.A.
DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF RAT TESTES: A METHOD FOR EARLY DETECTION OF ISCHEMIA MARKO KANGASNIEMI, ANTTI KAIPIA
AND
RAIMO JOENSUU
From the Department of Radiology, Helsinki University Central Hospital, Helsinki and Department of Urology, Tampere University Hospital, Tampere, Finland
ABSTRACT
Purpose: We investigated the feasibility of diffusion weighted magnetic resonance imaging (MRI) for the early detection of ischemia in the testis. Materials and Methods: Circulation to the right testis in Wistar rats was occluded by surgical ligation of the right funicle. The left side was sham operated and served as a control. The diffusion and T2-weighted MRI images of the 2 testes was performed postoperatively by a 1.5 Tesla MRI unit using a knee coil. On apparent diffusion coefficient images and T2-weighted images the region of interest values in the 2 testes were measured and statistically compared. Results: At 1 hour after testicular funicle ligation the apparent diffusion coefficient was 18% lower in the ischemic than in the sham operated testis (p ⬍0.0098). At 2 hours the difference was 20% (p ⬍0.0017). In the signal-to-noise ratio on T2-weighted images there was no difference in the left and right testes. Conclusions: Altered diffusion occurs in an ischemic testis, which can be measured on MRI at 1.5 Tesla. Thus, diffusion-weighted MRI may be a helpful method for the differential diagnosis of acute testicular torsion. KEY WORDS: testis; rats, Wistar; ischemia; magnetic; resonance imaging
Currently at most institutions ultrasonography is the primary imaging method of the acute scrotum. For diagnosing torsion of the testis sensitivity and specificity exceeding 90% have been reported for color Doppler ultrasound.1 However, acute scrotum in a clinical setting still represents a considerable diagnostic problem.2 Torsion of the appendix testis, epididymitis and testicular torsion often present with similar clinical symptoms. When color Doppler ultrasound cannot definitely rule out compromised perfusion, surgical exploration is often warranted. In most former studies imaging of the ischemic testis has been based on detecting reduced testicular blood flow. An imaging method based on detecting tissue ischemia instead of decreased perfusion would be ideal for imaging the acute scrotum because it would detect torsion of the testis at its early phase when some arterial perfusion is still present as well as an episode of incomplete testicular torsion. During the last decade diffusion-weighted magnetic resonance imaging (MRI) has become an important method for diagnosing acute stroke.3 The method is based on measuring the changes in cellular water content and in water diffusion through the cellular membrane, which occur at the early stages of tissue ischemia. In this study diffusion-weighted MRI was done to detect developing ischemia in testicular tissue. We examined whether diffusion is altered in hypoperfused testes at a level that can be measured by MRI at 1.5 Tesla at an early phase of ischemia. MATERIALS AND METHODS
Animals and surgical procedure. Six adult 5-month-old Wistar rats were used in this investigation. The Helsinki University Central Hospital animal care committee approved the experiments. Sagittal incisions (2 cm.) were made on each side of the scrotum while the animals were under anesthesia with fentanyl citrate at a dose of 0.25 mg./500 gm. animal weight and midatzolam at a dose of 0.25 mg./500 gm. animal weight. The
testes were exposed. A 3-zero polyglactin ligature was tightened around the right funicle to prevent perfusion to the right testis. The testes and funicle were placed back into the scrotum and the incision was sutured using a monofilament nylon suture. The left testis served as a sham operated control. After the completion of MRI at the end of each experiment the animals were sacrificed. The testes were removed and ischemia of the right testis was verified by inspection. The 2 testes were processed for histology, as previously described.4 Right and left testicular histology was compared under light microscopy. MRI of the testes. A 1.5 Tesla human MRI Magnetom Vision unit (Siemens, Erlangen, Germany) with a 25 mT/m maximum gradient capability and a standard knee coil was used to obtain all MRI images. A single acquisition of a conventional T2-weighed turbo spin-echo sequence (4,500/96) (repetition time in milliseconds/effective echo time in milliseconds) with an echo train length of 5 was performed with a matrix resolution of 210 ⫻ 256. The field of view was 210 mm. with a section thickness of 3 mm. and an intersection gap of 0.3 mm. Diffusion-weighted MRI was performed with the standard diffusion-weighted sequence (ep2d_d33a_100b1250_11.ekc). The diffusion sensitizing scheme contained 4 gradient and 2, 180-degree radio frequency pulses, and the diffusion gradients were activated in each of the 3 principal axes with gradient factor b values of 0 and 1,000 seconds per mm.2. Console software directly calculated the apparent diffusion coefficient maps. The end user controllable imaging parameters were repetition-to-echo time 4,000/100, field of view 210 mm., acquisition matrix 96 ⫻ 128, section thickness 3 mm. with 0.3 mm. intersection gap and 2 acquisitions. Data collection and analysis. The average signal of each testis was measured by a region of interest that covered the whole testicular section in each axial plane where the testis was visible or 3 to 5 planes per testis. The average of all values measured from the various planes represented the average signal value of that specific testis. Data were col-
Accepted for publication June 22, 2001. Supported by a research grant from Tampere University Hospital. 2542
DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF TESTIS
lected from apparent diffusion coefficient maps and T2weighted images 1 and 2 hours after surgical closure of the right funicle. The signal-to-noise ratio is reported as the testicular signal intensity level on T2-weighted images divided by background noise in root mean square. The 2-way Student’s t test was used to compare statistically the values calculated for the right testes with the values for the left testes. Apparent diffusion coefficient and testicular signal-to-noise ratio on T2-weighted MRI in right and left testes Time* (hrs.)
Mean Rt. Testis ⫾ SEM
Mean Lt. Testis ⫾ SEM
p Value
2543
RESULTS
The mean apparent diffusion coefficient was significantly lower in the right testes in which the funicle was ligated compared with the left testes. One hour after right funicle ligation the difference was 18% (p ⬍0.0098) and at 2 hours it was 20% (p ⬍0.0017, see table and fig. 1). The mean signalto-noise ratio measured on T2-weighted images was not significantly different in the right funicle ligated and left testes 1 or 2 hours after ligation (see table and fig. 2). Histological evaluation showed slight interstitial edema and blood congestion in the capillaries 1 hour after ligation but cellular morphology did not differ in the right and left testes (fig. 3). Thus, histological changes were considered to be less than grade 1 according to histological grading of testis torsion.5
Apparent diffusion coefficient 1 2
88 ⫾ 3 92 ⫾ 5
108 ⫾ 5 115 ⫾ 6
0.0098 0.0017
Signal-to-noise ratio 1 153 ⫾ 26 151 ⫾ 9 2 135 ⫾ 8 132 ⫾ 7 * Measured from the closing of the right testicular funicle.
FIG. 1. Axial caudocranial apparent diffusion coefficient map of testes of different rats. a, unoperated control animal. b and c, 1 hour after right funicle ligation reduction of signal intensity persisted in ischemic right testis. d and e, 2 hours after ligation reduction of signal intensity persisted in right testis.
DISCUSSION
To our knowledge we report the first use of diffusionweighted MRI to detect ischemic changes in the testis. The results show that a significant decrease in diffusion occurs in the hypoperfused testis and the resulting quantitative change in the apparent diffusion coefficient is sufficient to be measured using a 1.5 Tesla MRI unit. Furthermore, the decrease in diffusion is measurable at an early phase of ischemia before morphological changes develop in the testis. Currently ultrasonography is still the method of choice for
FIG. 2. Axial caudocranial T2-weighted image of rat testes. a, unoperated control rat. b, rat with ligated right funicle 1 hour after ligation. c, in rat with right funicle ligated no ischemia related change in testicular image was present after 2 hours.
2544
DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF TESTIS
the blood flow. Such a disturbance may well be missed by ultrasonography but if cellular ischemia occurs, diffusionweighted MRI may detect it. MRI by high resolution sequences other than diffusion also provide an accurate anatomical record of the target and, thus, other scrotal disorders such as testicular tumors or infarction can be reliably recognized.6, 7 Diffusion-weighted MRI may also have better reproducibility than ultrasonography. The information obtained does not depend as much on the experience or skills of the performing physician as with ultrasonography. Today diffusion-weighted MRI is widely performed to image ischemic changes in the brain, in which previous studies have demonstrated a signal change of up to 60% 1 hour after experimentally induced ischemia.8 The difference in the magnitude of the signal change in brain and testicular tissue may be due to the different cellular compositions and/or different tolerance of oxygen deprivation. Interestingly during testicular ischemia perfusion in the contralateral testis may also be reduced.9, 10 Thus, some decrease in the apparent diffusion coefficient may also have occurred in the sham operated testis in our series. In future studies a dedicated coil may be necessary to optimize the potential of diffusion-weighted MRI for detecting testicular ischemia. In conclusion, diffusion weighted MRI of the acute scrotum may become a rapid and helpful tool for diagnosing testicular torsion. REFERENCES
FIG. 3. Photomicrographs of rat testicular sections. a, unoperated control. b, 1 hour after testicular funicle ligation no ischemia related change was visualized in tissue or cellular morphology. Reduced from ⫻250.
imaging acute scrotum in the clinical setting. Although a high predictive value of ultrasonography has been reported for diagnosing torsion of the testis,1 in clinical studies the examination is occasionally inconclusive and torsion cannot be definitely ruled out. In such cases diffusion-weighted MRI may provide advantages compared with other imaging modalities. Instead of measuring blood flow this technique directly quantitates ischemia related changes within the cells.3 This finding may be useful because partial testicular torsion cause initially venous stasis instead of complete blockade of
1. Baker, L. A., Sigman, D., Mathews, R. I. et al: An analysis of clinical outcomes using color doppler testicular ultrasound for testicular torsion. Pediatrics, 105: 604, 2000 2. Dunne, P. J. and O’Loughlin, B. S.: Testicular torsion: time is the enemy. Aust N Z J Surg, 70: 441, 2000 3. Baird, A. E. and Warach, S.: Magnetic resonance imaging of acute stroke. J Cereb Blood Flow Metab, 18: 583, 1998 4. Kangasniemi, M., Kaipia, A., Toppari, J. et al: Cellular regulation of basal and FSH-stimulated cyclic AMP production in irradiated rat testes. Anat Rec, 227: 32, 1990 5. Mikuz, G.: Testicular torsion: simple grading for histological evaluation of tissue damage. Appl Pathol, 3: 134, 1985 6. Watanabe, Y., Dohke, M., Ohkubo, K. et al: Scrotal disorders: evaluation of testicular enhancement patterns at dynamic contrast-enhanced subtraction MR imaging. Radiology, 217: 219, 2000 7. Kodama, K., Yotsuyanagi, S., Fuse, H. et al: Magnetic resonance imaging to diagnose segmental testicular infarction. J Urol, 163: 910, 2000 8. Hoehn-Berlage, M., Norris, D. G., Kohno, K. et al: Evolution of regional changes in apparent diffusion coefficient during focal ischemia of rat brain: the relationship of quantitative diffusion NMR imaging to reduction in cerebral blood flow and metabolic disturbances. J Cereb Blood Flow Metab, 15: 1002, 1995 9. Nguyen, L., Lievano, G., Ghosh, L. et al: Effect of unilateral testicular torsion on blood flow and histology of contralateral testes. J Pediatr Surg, 34: 680, 1999 10. Salman, A. B., Mutlu, S., Iskit, A. B. et al: Hemodynamic monitoring of the contralateral testis during unilateral testicular torsion describes the mechanism of damage. Eur Urol, 33: 576, 1998
Vol. 166, 2542–2544, December 2001 Printed in U.S.A.
DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF RAT TESTES: A METHOD FOR EARLY DETECTION OF ISCHEMIA MARKO KANGASNIEMI, ANTTI KAIPIA
AND
RAIMO JOENSUU
From the Department of Radiology, Helsinki University Central Hospital, Helsinki and Department of Urology, Tampere University Hospital, Tampere, Finland
ABSTRACT
Purpose: We investigated the feasibility of diffusion weighted magnetic resonance imaging (MRI) for the early detection of ischemia in the testis. Materials and Methods: Circulation to the right testis in Wistar rats was occluded by surgical ligation of the right funicle. The left side was sham operated and served as a control. The diffusion and T2-weighted MRI images of the 2 testes was performed postoperatively by a 1.5 Tesla MRI unit using a knee coil. On apparent diffusion coefficient images and T2-weighted images the region of interest values in the 2 testes were measured and statistically compared. Results: At 1 hour after testicular funicle ligation the apparent diffusion coefficient was 18% lower in the ischemic than in the sham operated testis (p ⬍0.0098). At 2 hours the difference was 20% (p ⬍0.0017). In the signal-to-noise ratio on T2-weighted images there was no difference in the left and right testes. Conclusions: Altered diffusion occurs in an ischemic testis, which can be measured on MRI at 1.5 Tesla. Thus, diffusion-weighted MRI may be a helpful method for the differential diagnosis of acute testicular torsion. KEY WORDS: testis; rats, Wistar; ischemia; magnetic; resonance imaging
Currently at most institutions ultrasonography is the primary imaging method of the acute scrotum. For diagnosing torsion of the testis sensitivity and specificity exceeding 90% have been reported for color Doppler ultrasound.1 However, acute scrotum in a clinical setting still represents a considerable diagnostic problem.2 Torsion of the appendix testis, epididymitis and testicular torsion often present with similar clinical symptoms. When color Doppler ultrasound cannot definitely rule out compromised perfusion, surgical exploration is often warranted. In most former studies imaging of the ischemic testis has been based on detecting reduced testicular blood flow. An imaging method based on detecting tissue ischemia instead of decreased perfusion would be ideal for imaging the acute scrotum because it would detect torsion of the testis at its early phase when some arterial perfusion is still present as well as an episode of incomplete testicular torsion. During the last decade diffusion-weighted magnetic resonance imaging (MRI) has become an important method for diagnosing acute stroke.3 The method is based on measuring the changes in cellular water content and in water diffusion through the cellular membrane, which occur at the early stages of tissue ischemia. In this study diffusion-weighted MRI was done to detect developing ischemia in testicular tissue. We examined whether diffusion is altered in hypoperfused testes at a level that can be measured by MRI at 1.5 Tesla at an early phase of ischemia. MATERIALS AND METHODS
Animals and surgical procedure. Six adult 5-month-old Wistar rats were used in this investigation. The Helsinki University Central Hospital animal care committee approved the experiments. Sagittal incisions (2 cm.) were made on each side of the scrotum while the animals were under anesthesia with fentanyl citrate at a dose of 0.25 mg./500 gm. animal weight and midatzolam at a dose of 0.25 mg./500 gm. animal weight. The
testes were exposed. A 3-zero polyglactin ligature was tightened around the right funicle to prevent perfusion to the right testis. The testes and funicle were placed back into the scrotum and the incision was sutured using a monofilament nylon suture. The left testis served as a sham operated control. After the completion of MRI at the end of each experiment the animals were sacrificed. The testes were removed and ischemia of the right testis was verified by inspection. The 2 testes were processed for histology, as previously described.4 Right and left testicular histology was compared under light microscopy. MRI of the testes. A 1.5 Tesla human MRI Magnetom Vision unit (Siemens, Erlangen, Germany) with a 25 mT/m maximum gradient capability and a standard knee coil was used to obtain all MRI images. A single acquisition of a conventional T2-weighed turbo spin-echo sequence (4,500/96) (repetition time in milliseconds/effective echo time in milliseconds) with an echo train length of 5 was performed with a matrix resolution of 210 ⫻ 256. The field of view was 210 mm. with a section thickness of 3 mm. and an intersection gap of 0.3 mm. Diffusion-weighted MRI was performed with the standard diffusion-weighted sequence (ep2d_d33a_100b1250_11.ekc). The diffusion sensitizing scheme contained 4 gradient and 2, 180-degree radio frequency pulses, and the diffusion gradients were activated in each of the 3 principal axes with gradient factor b values of 0 and 1,000 seconds per mm.2. Console software directly calculated the apparent diffusion coefficient maps. The end user controllable imaging parameters were repetition-to-echo time 4,000/100, field of view 210 mm., acquisition matrix 96 ⫻ 128, section thickness 3 mm. with 0.3 mm. intersection gap and 2 acquisitions. Data collection and analysis. The average signal of each testis was measured by a region of interest that covered the whole testicular section in each axial plane where the testis was visible or 3 to 5 planes per testis. The average of all values measured from the various planes represented the average signal value of that specific testis. Data were col-
Accepted for publication June 22, 2001. Supported by a research grant from Tampere University Hospital. 2542
DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF TESTIS
lected from apparent diffusion coefficient maps and T2weighted images 1 and 2 hours after surgical closure of the right funicle. The signal-to-noise ratio is reported as the testicular signal intensity level on T2-weighted images divided by background noise in root mean square. The 2-way Student’s t test was used to compare statistically the values calculated for the right testes with the values for the left testes. Apparent diffusion coefficient and testicular signal-to-noise ratio on T2-weighted MRI in right and left testes Time* (hrs.)
Mean Rt. Testis ⫾ SEM
Mean Lt. Testis ⫾ SEM
p Value
2543
RESULTS
The mean apparent diffusion coefficient was significantly lower in the right testes in which the funicle was ligated compared with the left testes. One hour after right funicle ligation the difference was 18% (p ⬍0.0098) and at 2 hours it was 20% (p ⬍0.0017, see table and fig. 1). The mean signalto-noise ratio measured on T2-weighted images was not significantly different in the right funicle ligated and left testes 1 or 2 hours after ligation (see table and fig. 2). Histological evaluation showed slight interstitial edema and blood congestion in the capillaries 1 hour after ligation but cellular morphology did not differ in the right and left testes (fig. 3). Thus, histological changes were considered to be less than grade 1 according to histological grading of testis torsion.5
Apparent diffusion coefficient 1 2
88 ⫾ 3 92 ⫾ 5
108 ⫾ 5 115 ⫾ 6
0.0098 0.0017
Signal-to-noise ratio 1 153 ⫾ 26 151 ⫾ 9 2 135 ⫾ 8 132 ⫾ 7 * Measured from the closing of the right testicular funicle.
FIG. 1. Axial caudocranial apparent diffusion coefficient map of testes of different rats. a, unoperated control animal. b and c, 1 hour after right funicle ligation reduction of signal intensity persisted in ischemic right testis. d and e, 2 hours after ligation reduction of signal intensity persisted in right testis.
DISCUSSION
To our knowledge we report the first use of diffusionweighted MRI to detect ischemic changes in the testis. The results show that a significant decrease in diffusion occurs in the hypoperfused testis and the resulting quantitative change in the apparent diffusion coefficient is sufficient to be measured using a 1.5 Tesla MRI unit. Furthermore, the decrease in diffusion is measurable at an early phase of ischemia before morphological changes develop in the testis. Currently ultrasonography is still the method of choice for
FIG. 2. Axial caudocranial T2-weighted image of rat testes. a, unoperated control rat. b, rat with ligated right funicle 1 hour after ligation. c, in rat with right funicle ligated no ischemia related change in testicular image was present after 2 hours.
2544
DIFFUSION WEIGHTED MAGNETIC RESONANCE IMAGING OF TESTIS
the blood flow. Such a disturbance may well be missed by ultrasonography but if cellular ischemia occurs, diffusionweighted MRI may detect it. MRI by high resolution sequences other than diffusion also provide an accurate anatomical record of the target and, thus, other scrotal disorders such as testicular tumors or infarction can be reliably recognized.6, 7 Diffusion-weighted MRI may also have better reproducibility than ultrasonography. The information obtained does not depend as much on the experience or skills of the performing physician as with ultrasonography. Today diffusion-weighted MRI is widely performed to image ischemic changes in the brain, in which previous studies have demonstrated a signal change of up to 60% 1 hour after experimentally induced ischemia.8 The difference in the magnitude of the signal change in brain and testicular tissue may be due to the different cellular compositions and/or different tolerance of oxygen deprivation. Interestingly during testicular ischemia perfusion in the contralateral testis may also be reduced.9, 10 Thus, some decrease in the apparent diffusion coefficient may also have occurred in the sham operated testis in our series. In future studies a dedicated coil may be necessary to optimize the potential of diffusion-weighted MRI for detecting testicular ischemia. In conclusion, diffusion weighted MRI of the acute scrotum may become a rapid and helpful tool for diagnosing testicular torsion. REFERENCES
FIG. 3. Photomicrographs of rat testicular sections. a, unoperated control. b, 1 hour after testicular funicle ligation no ischemia related change was visualized in tissue or cellular morphology. Reduced from ⫻250.
imaging acute scrotum in the clinical setting. Although a high predictive value of ultrasonography has been reported for diagnosing torsion of the testis,1 in clinical studies the examination is occasionally inconclusive and torsion cannot be definitely ruled out. In such cases diffusion-weighted MRI may provide advantages compared with other imaging modalities. Instead of measuring blood flow this technique directly quantitates ischemia related changes within the cells.3 This finding may be useful because partial testicular torsion cause initially venous stasis instead of complete blockade of
1. Baker, L. A., Sigman, D., Mathews, R. I. et al: An analysis of clinical outcomes using color doppler testicular ultrasound for testicular torsion. Pediatrics, 105: 604, 2000 2. Dunne, P. J. and O’Loughlin, B. S.: Testicular torsion: time is the enemy. Aust N Z J Surg, 70: 441, 2000 3. Baird, A. E. and Warach, S.: Magnetic resonance imaging of acute stroke. J Cereb Blood Flow Metab, 18: 583, 1998 4. Kangasniemi, M., Kaipia, A., Toppari, J. et al: Cellular regulation of basal and FSH-stimulated cyclic AMP production in irradiated rat testes. Anat Rec, 227: 32, 1990 5. Mikuz, G.: Testicular torsion: simple grading for histological evaluation of tissue damage. Appl Pathol, 3: 134, 1985 6. Watanabe, Y., Dohke, M., Ohkubo, K. et al: Scrotal disorders: evaluation of testicular enhancement patterns at dynamic contrast-enhanced subtraction MR imaging. Radiology, 217: 219, 2000 7. Kodama, K., Yotsuyanagi, S., Fuse, H. et al: Magnetic resonance imaging to diagnose segmental testicular infarction. J Urol, 163: 910, 2000 8. Hoehn-Berlage, M., Norris, D. G., Kohno, K. et al: Evolution of regional changes in apparent diffusion coefficient during focal ischemia of rat brain: the relationship of quantitative diffusion NMR imaging to reduction in cerebral blood flow and metabolic disturbances. J Cereb Blood Flow Metab, 15: 1002, 1995 9. Nguyen, L., Lievano, G., Ghosh, L. et al: Effect of unilateral testicular torsion on blood flow and histology of contralateral testes. J Pediatr Surg, 34: 680, 1999 10. Salman, A. B., Mutlu, S., Iskit, A. B. et al: Hemodynamic monitoring of the contralateral testis during unilateral testicular torsion describes the mechanism of damage. Eur Urol, 33: 576, 1998