Faculty Bibliography

INTRODUCTION/BACKGROUND:There has been limited attention to pathological features of basilar artery atherosclerosis. It has been assumed that pathology of basilar artery atherosclerosis mimics that of other vascular beds. METHODS:To define the nature of the basilar artery atherosclerotic lesions, we analyzed postmortem intracranial artery samples from eight subjects with history of stroke. RESULTS:Atherosclerotic lesions were present in 7/8 arteries examined, with a mean estimated stenosis of 34%. Lumen thrombus with a disrupted fibrous cap was seen in 1 lesion; the remaining 6 lesions revealed a thick fibrous cap. Neovascularity and calcification were seen in 1 lesion and mild to moderate inflammation was seen in 3 lesions. Necrotic core was present in 4/7 lesions, and was associated with plaque rupture in the only disrupted lesion. CONCLUSIONS:Basilar artery atherosclerotic lesions were relatively benign in this series of patients presenting with stroke. While confirmation is needed with larger sample size, the relative paucity of neovascularity suggests a possibly distinctive histopathological profile.

PURPOSE/OBJECTIVE:To our knowledge the optimal freeze cycle length in renal cryotherapy is unknown. Ten-minute time based freeze cycles were compared to temperature based freeze cycles to -20C. MATERIALS AND METHODS/METHODS:Laparoscopic renal cryotherapy was performed on 16 swine. Time based trials consisted of a double 10-minute freeze separated by a 5-minute thaw. Temperature based trials were double cycles of 1, 5 or 10-minute freeze initiated after 1 of 4 sensors indicated -20C. A 5-minute active thaw was used between freeze cycles. Control trials consisted of cryoneedle placement for 25 minutes without freeze or thaw. Viability staining and histological analysis were done. RESULTS:There was no difference in cellular necrosis between any of the temperature based freeze cycles (p = 0.1). Time based freeze cycles showed more nuclear pyknosis, indicative of necrosis, than the 3 experimental freeze cycles for the renal cortex (p = 0.05) but not for the renal medulla (p = 0.61). Mean time to -20C for freeze cycle 1 was 19 minutes 10 seconds (range 9 to 46 minutes). In 4 of 21 trials (19%) -20C was never attained despite freezing for 25 to 63 minutes. CONCLUSIONS:There was no difference in immediate cellular necrosis among double 1, 5 or 10-minute freeze cycles. Cellular necrosis was evident on histological analysis for trials in which -20C was attained and in freeze cycles based on time alone. With a standard 10-minute cryoablation period most treated parenchyma 1 cm from the probe never attained -20C. Cell death appeared to occur at temperatures warmer than -20C during renal cryotherapy.

OBJECTIVES/OBJECTIVE:To evaluate the immunohistochemical profile of a carcinoid (low grade neuroendocrine tumor of the kidney) from a patient with lymph node positive disease who remains disease free for 31 months after radical nephrectomy, lymph node dissection, and adjuvant therapy with sunitinib malate. METHODS:Immunohistochemical staining was performed for chromogranin, synaptophysin, CD31, VEGF, HIF-1α, HIF-2, and Glut-1. Staining was evaluated in 3 high-power fields and samples scored as strongly positive (3+), moderately positive (2+), weakly positive (1+), or negative (0). A clear cell renal cell carcinoma was used as positive control. RESULTS:Immunohistochemical staining was strongly positive VEGF, weak to moderately positive for HIF-2, and negative for HIF-1α and Glut-1. CONCLUSIONS:Our case of primary renal carcinoid stained intensely for VEGF and HIF-2, consistent with a VHL-HIF1-HIF2-Glut1 independent pathway for VEGF activation. These data suggest that like other neuroendocrine tumors, primary renal carcinoid is a potential target for anti-angiogenic therapy with sunitinib.

OBJECTIVES/OBJECTIVE:Technetium-99m-labeled matrix metalloproteinase inhibitor (MPI) was used for the noninvasive assessment of matrix metalloproteinase (MMP) activity in atherosclerotic plaques after minocycline (MC) intervention. BACKGROUND:MMP activity in atherosclerosis contributes to plaque instability. Some antimicrobial agents may attenuate MMP activity. METHODS:Atherosclerotic lesions were produced in 38 rabbits with a high cholesterol diet for 4 months; 5 groups of rabbits, in the fourth month, received fluvastatin (FS) (n = 6), low-dose MC (n = 7), high-dose MC (n = 7), a combination of low-dose MC and FS (n = 6), or no intervention (n = 12); 8 unmanipulated rabbits were used as disease controls. Micro-single-photon emission computed tomography imaging was performed in all animals after intravenous MPI administration, followed by pathologic characterization of the aorta. A cell culture study evaluated the effect of MC on MMP production by activated human monocytes. RESULTS:MPI uptake was visualized best in untreated atherosclerotic animals (percent injected dose per gram MPI uptake, 0.11 +/- 0.04%). MPI uptake was reduced in the FS (0.06 +/- 0.01%; p < 0.0001), high-dose MC (0.05 +/- 0.01%; p < 0.0001), and MC-FS (0.05 +/- 0.005%; p < 0.0001) groups. Low-dose MC did not resolve MPI uptake significantly (0.08 +/- 0.02; p = 0.167). There was no incremental benefit of the combination of MC and FS. MPI uptake showed a significant correlation with plaque MMP-2, and MMP-9 activity. MMP-9 release from tumor necrosis factor-alpha-activated macrophages was abrogated by incubation with MC. CONCLUSIONS:Molecular imaging of MMP activity in atherosclerotic plaque allows for the study of the efficacy of therapeutic interventions. MC administration resulted in substantial reduction in plaque MMP activity and histologically verified plaque stabilization. MC was found to be equally effective as FS.

UNLABELLED:Ischemic insult to the myocardium is associated with cardiomyocyte apoptosis. Because apoptotic cell death is characterized by phosphatidylserine externalization on cell membrane and annexin-A5 (AA5) avidly binds to phosphatidylserine, we hypothesized that radiolabeled AA5 should be able to identify the regions of myocardial ischemia. METHODS:Models of brief myocardial ischemia by the occlusion of the coronary artery for 10 min (I-10) and reperfusion for 180 min (R-180) for the detection of phosphatidylserine exteriorization using (99m)Tc-labeled AA5 and gamma-imaging were produced in rabbits. (99m)Tc-AA5 uptake after brief ischemia was compared with an I-40/R-180 infarct model. Histologic characterization of both myocardial necrosis and apoptosis was performed in ischemia and infarct models. Phosphatidylserine exteriorization was also studied in a mouse model, and the dynamics and kinetics of phosphatidylserine exposure were assessed using unlabeled recombinant AA5 and AA5 labeled with biotin, Oregon Green, or Alexa 568. Appropriate controls were established. RESULTS:Phosphatidylserine exposure after ischemia in the rabbit heart could be detected by radionuclide imaging with (99m)Tc-AA5. Pathologic characterization of the explanted rabbit hearts did not show apoptosis or necrosis. Homogenization and ultracentrifugation of the ischemic myocardial tissue from rabbit hearts recovered two thirds of the radiolabeled AA5 from the cytoplasmic compartment. Murine experiments demonstrated that the cardiomyocytes expressed phosphatidylserine on their cell surface after an ischemic insult of 5 min. Phosphatidylserine exposure occurred continuously for at least 6 h after solitary ischemic insult. AA5 targeted the exposed phosphatidylserine on cardiomyocytes; AA5 was internalized into cytoplasmic vesicles within 10-30 min. Twenty-four hours after ischemia, cardiomyocytes with internalized AA5 had restored phosphatidylserine asymmetry of the sarcolemma, and no detectable phosphatidylserine remained on the cell surface. The preadministration of a pan-caspase inhibitor, zVAD-fmk, prevented phosphatidylserine exposure after ischemia. CONCLUSIONS:After a single episode of ischemia, cardiomyocytes express phosphatidylserine, which is amenable to targeting by AA5, for at least 6 h. Phosphatidylserine exposure is transient and internalized in cytoplasmic vesicles after AA5 binding, indicating the reversibility of the apoptotic process.

Chronic mucosal and submucosal injury can lead to persistent inflammation and tissue remodeling. We hypothesized that microstructural and mechanical properties of the airway wall could be derived from multiphoton images. New Zealand White rabbits were intubated, and the tracheal epithelium gently denuded every other day for five days (three injuries). Three days following the last injury, the tracheas were excised for multiphoton imaging, mechanical compression testing, and histological analysis. Multiphoton imaging and histology confirm epithelial denudation, mucosal ulceration, subepithelial thickening, collagen deposition, immune cell infiltration, and a disrupted elastin network. Elastase removes the elastin network and relaxes the collagen network. Purified collagenase removes epithelium with subtle subepithelial changes. Young's modulus [(E) measured in kiloPascal] was significantly elevated for the scrape injured (9.0+/-3.2) trachea, and both collagenase (2.6+/-0.4) and elastase (0.8+/-0.3) treatment significantly reduced E relative to control (4.1+/-0.7). E correlates strongly with second harmonic generation (SHG) signal depth decay for enzyme-treated and control tracheas (R(2)=0.77), but not with scrape-injured tracheas. We conclude that E of subepithelial connective tissue increases on repeated epithelial wounding, due in part to changes in elastin and collagen microstructure and concentration. SHG depth decay is sensitive to changes in extracellular matrix content and correlates with bulk Young's modulus.

PURPOSE/OBJECTIVE:Adipose tissue has been suggested to contribute to the pathogenesis of various disease states, including prostate cancer. We investigated the association of cytokines and growth factors secreted by periprostatic adipose tissue with pathological features of aggressive prostate cancer. MATERIALS AND METHODS/METHODS:Periprostatic adipose tissue was harvested from patients undergoing radical prostatectomy and cultured for 24 hours to generate conditioned medium or snap frozen immediately for functional signaling profiling. Multiplex analysis of the periprostatic adipose tissue conditioned medium was used to detect cytokine levels and compared to patient matched serum from 7 patients. Interleukin-6 in serum and periprostatic adipose tissue conditioned medium was further analyzed by enzyme-linked immunosorbent assay and correlated with clinical variables, such as age, body mass index and Gleason score, in 45 patients. Interleukin-6 expression in periprostatic adipose tissue was determined by immunohistochemistry. Reverse phase protein microarray technology was used to analyze cell signaling networks in periprostatic adipose tissue. RESULTS:Interleukin-6 in periprostatic adipose tissue conditioned medium was approximately 375 times greater than that in patient matched serum and levels correlated with pathological grade. This finding was further extended by cell signaling analysis of periprostatic adipose tissue, which showed greater phosphorylation on Stat3 with high grade tumors (any component of Gleason score 4 or 5). CONCLUSIONS:Higher Gleason score correlated with high levels of conditioned medium derived interleukin-6. Moreover, cell signaling analysis of periprostatic adipose tissue identified activated signaling molecules, including STAT3, that correlated with Gleason score. Since STAT3 is interleukin-6 regulated, these findings suggest that periprostatic adipose tissue may have a role in modulating prostate cancer aggressiveness by serving as a source of interleukin-6. Also, we found low numbers of inflammatory cells in the fat, suggesting that adipocytes are the major secretors of interleukin-6.

PURPOSE/OBJECTIVE:Concern exists over a lack of tactile sensation and positive surgical margins (PSMs) in patients undergoing robot-assisted radical prostatectomy. We report our PSM rates in our most current 500 cases and particularly in clinically high-risk disease. MATERIALS AND METHODS/METHODS:After implementation of our present technique at case #251, we report PSM rates according to pathologic stage and D'Amico's risk stratification: low risk (prostate-specific antigen [PSA] <10, Gleason score [GS] 5-6, cT1-T2A), intermediate risk (PSA 10-20, GS 7, cT2B), and high risk (PSA >20, GS 8-10, cT3). Patients with cT2b/T3 disease or GS 8 to 10 and multiple cores with >30% involvement underwent wide excision of the neurovascular bundle. PSM was defined as ink on tumor. RESULTS:The overall PSM rate was 7.4%: pT2 = 3.1%, pT3 = 15.9%, and pT4 = 55.6%. PSMs occurred in 13 (4.9%) low, 10 (5.8%) intermediate, and 14 (22.6%) high D'Amico risk patients. Of the 62 high-risk patients, the median PSA was 6.9 (range 2.2-97.9); biopsy GS was 6 to 7 (26%) and 8 to 10 (74%). For preoperatively palpable disease, the PSM rate was 9.9%: cT1 = 6.0%, cT2 = 7.7%, and cT3 = 26.3%. No PSMs occurred along the neurovascular bundle. CONCLUSION/CONCLUSIONS:Since 2005, 500 men with clinically low-, intermediate-, and high-risk prostate cancer have undergone robot-assisted radical prostatectomy with acceptable surgical margin rates. In patients with high-risk and usually palpable disease, PSM rates were also acceptable despite the lack of tactile sensation with the robot.

BACKGROUND:Macrophage apoptosis and MMP activity contribute to vulnerability of atherosclerotic plaques to rupture. By employing molecular imaging techniques, we investigated if apoptosis and MMP release are interlinked. METHODS:Atherosclerosis was produced in rabbits receiving high-cholesterol diet (HC), who underwent dual radionuclide imaging with (99m)Tc-labeled matrix metalloproteinase inhibitor (MPI) and (111)In-labeled annexin A5 (AA5) using micro-SPECT/CT. %ID/g MPI and AA5 uptake was measured, followed by histological characterization. Unmanipulated animals were used as disease controls. Correlation between MPI and AA5 uptake was undertaken and relationship confirmed in culture study of activated THP-1 monocytes. RESULTS:MPI and AA5 uptake was best visualized in HC diet animals (n = 6) and reduced significantly after fluvastatin treatment (n = 4) or diet withdrawal (n = 3). %ID/g MPI (.087 +/- .018%) and AA5 (.03 +/- .01%) uptake was higher in HC than control (n = 6) animals (.014 +/- .004%, P < .0001; .0007 +/- .0002%, P < .0001), and reduced substantially after diet or statin intervention. There was a significant correlation between MPI and AA5 uptake (r = .62, P < .0001), both correlated with pathologically verified MMP-9 activity, macrophage content, and TUNEL staining. In vitro studies demonstrated MMP-9 release in culture medium from apoptotic THP-1 monocytes. CONCLUSIONS:The present study suggests that apoptosis and MMP are interrelated in atherosclerotic lesions and the targeting of more than one molecular candidate is feasible by molecular imaging.

Diffusion-weighted imaging has been widely accepted as a powerful imaging technique in neuroradiology. Until recently, the inclusion of diffusion-weighted sequences in body imaging protocols has been hindered by technical limitations. However, with advances in magnetic resonance (MR) imaging technology and technique, these limitations are being overcome. The addition of diffusion-weighted sequences to routine abdominopelvic MR imaging protocols has been found to yield diagnostically useful information with only a minimal increase in imaging time. More specifically, the use of diffusion-weighted imaging in the genitourinary system can facilitate the detection and characterization of genitourinary tract lesions that demonstrate equivocal signal intensity characteristics with routine MR imaging sequences. Diffusion-weighted imaging is not only helpful in differentiating benign from malignant processes, but it can also be used to assess meta-static lesions, possible tumor recurrence, and treatment response. Because it does not require injection of a gadolinium-based contrast agent, diffusion-weighted imaging can be used in patients with renal insufficiency or contrast material allergy. Most of the body diffusion-weighted imaging studies reported in the literature to date have been conducted with 1.5-T magnets. However, the feasibility of body diffusion-weighted imaging at 3.0 T is currently under investigation in an effort to determine the efficacy of the routine inclusion of diffusion-weighted imaging sequences in 3.0-T body MR imaging protocols.