James E. Coad

Cryosurgical changes in the porcine kidney: histologic analysis with thermal history correlation

Advances in minimally invasive renal cryosurgery have renewed interest in the relative contributions of direct cryothermic and secondary vascular injury-associated ischemic cell injury. Prior studies have evaluated renal cryolesions seven or more days post-ablation and postulated that vascular injury is the primary cell injury mechanism; however, the contributions of direct versus secondary cell injury are not morphologically distinguishable during the healing/repair stage of a cryolesion. While more optimal to evaluate this issue, minimal acute (< or = 3 days) post-ablation histologic data with thermal history correlation exists. This study evaluates three groups of porcine renal cryolesions: Group (1) in vitro non-perfused (n = 5); Group (2) in vivo 2-h post-ablation perfused (n = 5); and Group (3) in vivo 3-day post-ablation perfused (n = 6). The 3.4 mm argon-cooled cryoprobes thermal history included a 75 degrees C/min cooling rate, -130 degrees C end temperature, 60 degrees C/min thawing rate, and 15-min freeze time. An enthalpy-based mathematical model with a 2-D transient axisymmetric numerical solution with blood flow consideration was used to determine the thermal history within the ice ball. All three groups of cryolesions showed histologically similar central regions of complete cell death (CD) and transition zones of incomplete cell death (TZ). The CD had radii of 1.4, 1.1, and 1.0 cm in the non-perfused, 2-h and 3-day lesions, respectively. Capillary thrombosis was present in the 2-h perfused cryolesions with the addition of TZ arteriolar/venous thrombosis in the 3-day perfused lesions. Thermal modeling revealed the outer CD boundary in all three groups experienced similar thermal histories with an approximately -20 degrees C end temperature and 2 degrees C/min cooling and thawing rates. The presence of similar CD histology and in vitro/in vivo thermal histories in each group suggests that direct cryothermic cell injury, prior to or synchronous with vascular thrombosis, is a primary mediator of cell death in renal cryolesions.

In Vitro Assessment of the Efficacy of Thermal Therapy in Human Benign Prostatic Hyperplasia

The successful management of BPH with minimally invasive thermal therapies requires a firm understanding of the temperature–time relationship for tissue destruction. In order to accomplish this objective, the present in vitro study assesses the cellular viability of human BPH tissue subjected to an experimental matrix of different temperature–time combinations. Hyperplastic prostate tissue was obtained from 10 radical prostatectomy specimens resected for adenocarcinoma. A portion of hyperplastic tissue from the lateral lobe of each prostate was sectioned into multiple 1u2009mm thick tissue strips, placed on a coverslip and thermally treated on a controlled temperature copper block with various temperatures (45–70°C) for various times (1–60u2009min). After heat treatment, the tissue slices were cultured for 72u2009h and viability was assessed using two independent assays: histology and dye uptake for stromal tissue and using histology alone for the glandular tissue. The hyperplastic human prostate tissue showed a progressive histological increase in irreversible injury with increasing temperature–time severity. The dye uptake and histology results for stromal viability were similar for all temperature–time combinations. In vitro thermal injury showed 85–90% stromal destruction (raw data) of human BPH for temperature–time combinations of 45°C for 60u2009min, 50°C for 30u2009min, 55°C for 5u2009min, 60°C for 2u2009min and 70°C for 1u2009min. Apoptosis was also identified in the control and milder treated tissues with the degree of glandular apoptosis (about 20%) more than that seen in the stromal regions (<5%). The Arrhenius model of injury was fitted to the data for conditions leading to a 90% drop in viability (normalized to control) obtained for stromal tissue. The activation energies (E) were 40.1 and 38.4u2009kcal/mole for the dye uptake study and histology, respectively, and the corresponding frequency factors (A) were 1.1 × 1024 and 7.78 × 1022/s. This study presents the first temperature–time versus tissue destruction relation for human BPH tissue. Moreover, it supports the concept that higher temperatures can be used for shorter durations to induce tissue injury comparable with the current clinically recommended lower temperature–longer time treatments (i.e. 45°C for 60u2009min) for transurethral microwave thermotherapy of the prostate.

Correlation of PCR-detected clonal gene rearrangements with bone marrow morphology in patients with B-lineage lymphomas.

Bone marrow biopsy is the conventional staging and posttherapy evaluation method for assessing marrow involvement by lymphoma. Polymerase chain reactions (PCR) for antigen receptor rearrangements have the potential to increase the detection of minimal degrees of marrow involvement. The present study is a concurrent morphologic and PCR evaluation of 225 staging or posttherapy marrow biopsies from 127 patients with B-lineage non-Hodgkins lymphoma. The biopsies were morphologically categorized into four groups: group 1 (positive for lymphoma), 60 biopsies (27%); group 2 (suspicious for lymphoma), 20 biopsies (9%); group 3 (lymphocytic lesions of indeterminate biology), 22 biopsies (10%); and group 4 (negative for lymphoma), 123 biopsies (54%). Molecular studies were performed on concurrently obtained aspirates and used consensus immunoglobulin-heavy-chain (IgH) and IgH/bcl-2 gene PCR primers. A molecular clone was detected in 53 of the 225 aspirates (24%): group 1, 34 aspirates (57%); group 2, five aspirates (25%); group 3, one aspirate (5%); and group 4, 13 aspirates (11%). A PCR-positive aspirate was present in 47% of follicular lymphomas, 58% of diffuse large cell lymphomas, and 72% of the other lymphomas in the group I specimens. Morphology or PCR was positive in 79 of the 225 cases (35%). The molecular detection of clonality in the aspirate DNA from cases with positive morphologic findings was lower than anticipated. The discordance between morphology and PCR results may be related to sample variation between the trephine biopsy and aspirate, a failure to aspirate sufficient lymphoma cells, or insufficient primer homology for amplification. DNA extracted from trephine sections may provide results more concordant with morphology, because PCR detected a clone in 10 of 11 DNA specimens extracted from trephine biopsies with positive morphologic findings and PCR negative aspirates.

In vitro thermal therapy of AT-1 Dunning prostate tumours

To advance the utility of prostate thermal therapy, this study investigated the thermal thresholds (temperature-time) for prostate tissue destructionin vitro. The AT-1 Dunning prostate tumour model was chosen for the study. Three hundred micron thick sections were subjected to controlled temperature-time heating, which ranged from low (40°C, 15u2009min) to high thermal exposures (70°C, 2u2009min) (n = 6). After subsequent tissue culture at 37°C, the sections were evaluated for tissue injury at 3, 24 and 72u2009h by two independent methods: histology and dye uptake. A graded increase in injury was identified between the low and high thermal exposures. Maximum histologic injury occurred above 70°C, 1u2009min with >95% of the tissue area undergoing significant cell injury and coagulative necrosis. The control and 40°C, 15u2009min sections showed histologic evidence of apoptosis following 24 and 72u2009h in culture. Similar signs of apoptosis were minimal or absent at higher thermal histories. Vital-dye uptake quantitatively confirmed complete cell death after 70°C, 2u2009min. Using the dye data, Arrhenius analysis showed an apparent breakpoint at 50°C, with activation energies of 135.8u2009kcal/mole below and 4.7u2009kcal/mole above the threshold after 3u2009h in culture. These results can be used as a conservative benchmark for thermal injury in the cancerous prostate. Further characterization of the response to thermal therapy in an animal model and in human tissues will be important in establishing the efficacy of the procedure

Splenectomy in Lymphoproliferative Disorders: a Report on 70 Cases and Review of the Literature

Between February, 1970 and September, 1991, we performed splenectomies on 70 patients with chronic lymphoproliferative disorders including primary leukemias: 19 B-cell chronic lymphocytic leukemia, 1 B-cell prolymphocytic leukemia, 22 hairy cell leukemias, 4 large granular lymphocytic leukemias, 1 T-cell prolymphocytic leukemia, and non-Hodgkins lymphomas (NHL): 10 splenic lymphomas with villous lymphocytes, 4 follicular lymphomas, 5 mantle cell lymphomas, 3 lymphoplasmacytic and 1 large cell NHL. The primary indications for surgery in this series were therapy-resistant disease (40%) and therapeutic splenectomy (38%). Postsplenectomy, 70% of patients had a complete hematological response, 23% had a partial response, and 7% were nonresponsive. Median treatment-free survival correlated with the hematologic response postsplenectomy and the underlying diagnosis. Better treatment-free survivals were seen in patients with lesser degrees of anemia and thrombocytopenia. Overall, improvements were more pronounced in the B-cell than in the T-cell disorders. Indications for further therapy, postoperative morbidity and mortality, and survival times are discussed along with a review of the literature. These findings advocate a continuing role for splenectomy in symptomatic lymphoid malignancies running with splenomegaly and hypersplenism.

Posttransplant lymphoproliferative disorder in pancreas transplantation: a single-center experience.

Background. Posttransplant lymphoproliferative disorder (PTLD) is a rare, serious complication of transplantation. The characteristics and associations of this disease in pancreas recipients have not been extensively studied. Methods. From January 1988 through December 2002, 787 pancreas and 569 kidney-pancreas transplants were performed at our institution. Eighteen pancreas recipients developed polymorphic PTLD or malignant lymphoma. Data on clinical course, organ involvement, molecular characteristics, and association with immunosuppression and recent cytomegalovirus (CMV) infection were compiled from the institutional transplant database. Patient survival was compared to recipients of liver and kidney transplants at the same center by using Kaplan-Meier analysis. Results. The 5-year cumulative incidence of PTLD in simultaneous pancreas-kidney, pancreas after kidney, and pancreas transplant alone recipients was 2.5%, 1.2%, and 1.0%, respectively (P = 0.23). A noticeably, but not significantly, higher cumulative incidence was seen in the more recent era since 1995 (2.1% vs. 0.9%, P = 0.15). PTLD in pancreas recipients carried a worse prognosis than in liver or kidney for recipients B-cell, early-onset, and Epstein Barr virus-positive lesions. PTLD was more aggressive in pancreas recipients, with a higher stage at presentation and a trend to more bone marrow involvement. There appeared to be a tendency toward association with recent CMV infection. Since 1995, PTLD recipients have had a lower exposure to antilymphocyte preparations (25 ± 5 vs. 10 ± 0.8)(P < 0.05). Conclusions. PTLD in pancreas recipients remains a rare but aggressive disease, and carries a worse prognosis in comparison to other transplant recipients. These heavily immunosuppressed patients, who often face multiple transplants, may be at greater risk; CMV infection may play an antecedent role.

Transplacental transfer of small-cell carcinoma of the lung.

To the Editor: Although the estimated rate of cancer during pregnancy is 1 case per 1000 live births,1 and placental metastases are not uncommon, transplacental transmission of maternal tumors is r...

Cryothermic and Hyperthermic Treatments of Human Leiomyomata and Adjacent Myometrium and Their Implications for Laparoscopic Surgery

STUDY OBJECTIVEnTo evaluate the effects and feasibility of direct cryothermic and hyperthermic therapy on leiomyomata and adjacent myometrium, and to contribute to evidence-based treatment thresholds based on measurements of direct cell injury.nnnDESIGNnExperimental study (Canadian Task Force classification II-2).nnnSETTINGnUniversity hospital.nnnSUBJECTSnLeiomyoma and myometrium tissue from 10 women undergoing total abdominal hysterectomy with or without bilateral salpingo-oophorectomy.nnnINTERVENTIONnIn vitro cryothermic or hyperthermic therapy was performed with representative leiomyoma and myometrium tissue samples. Using a directional solidification stage to simulate cryothermic therapy, 10 leiomyoma and 6 myometrium specimens were cooled in vitro at a rate of -5 degrees C/minute to end temperatures of -20 degrees, -40 degrees, -60 degrees, and -80 degrees C with a 15-minute hold period and then rapidly thawed to 21 degrees C. Hyperthermic therapy was simulated using a preheated 45 degrees, 55 degrees, 60 degrees, 65 degrees, 70 degrees, 75 degrees, and 80 degrees C constant temperature copper heating block with a 10-minute treatment period. In conjunction with tissue culturing and control tissues, cell death was assessed with routine histology and viability dyes (ethidium homodimer/Hoechst).nnnMEASUREMENTS AND MAIN RESULTSnIn cryothermic results, leiomyomata cell death (LCD) increased from 12% to 27% by histology and 26% to 38% by viability dye assay over the thermal range from -20 degrees to -80 degrees C, respectively. Myometrial cell death (MCD) increased from 10% to 12% and 4% to 20% for the same measurements, respectively. Whereas MCD appeared relatively stable from -40 degrees to -80 degrees C, it was significantly less than LCD over this range (p <0.05). For hyperthermic results, LCD increased from 17% to 88% by histology with progressive temperature increase from 45 degrees to 80 degrees C, respectively. The MCD showed a similar increase from 16% to 91% by histology over this temperature range. Hyperthermic histology and dye assay results were similar for LCD and MCD.nnnCONCLUSIONSnIn comparison with myometrium, leiomyomata showed greater direct cryothermic and equal hyperthermic cell injury. Whereas cell death increased up to 70 degrees C and down to -80 degrees C, the interval increases in cell injury diminished with more extreme temperatures. In vivo studies of combined direct and ischemic vascular injury thresholds have yet to be performed, but direct LCD matrixes determined in this study will help provide guidelines for minimally invasive surgical techniques for the treatment of leiomyomata.

Lesion formation and visualization using dual-mode ultrasound phased arrays

A 1 MHz spherical-section 64-element linear piezo-composite phased array and a supporting driving and data acquisition system were recently tested for use as a dual-mode high intensity focused ultrasound (HIFU) applicator system. In therapeutic mode, the array was shown to be capable of producing focal intensities in excess of 3500 W/cm/sup 2/ at its geometric center (100 mm radius of curvature). Imaging tests of quality assurance tissue-mimicking phantoms as well as computer simulations confirmed that the array has an oval-shaped imaging field of view (1xFOV) with nearly 50 dB, dynamic range extending from 70 to 120 mm in the axial direction and /spl plusmn/ 20 mm in the transverse direction. This is larger than its therapeutic operating field, which is defined as the region around the geometric focus where the loss in intensity gain is within 1 dB from the intensity gain at the geometric center. We have tested the array imaging capability in the visualization of the formation of discrete and volumetric lesions in freshly excised porcine liver samples. Experimental results clearly show that the enhancement of the tissue echogenicity at the lesion location is more pronounced in the harmonic images when compared to images at the fundamental. Results also clearly show that the harmonic imaging mode produces more accurate mapping of the lesion size and shape as determined by histologic evaluation.

Molecular Assessment of Clonality in Lymphoproliferative Disorders: I. Immunoglobulin Gene Rearrangements.

The non-Hodgkins lymphomas are a diverse group of malignancies with characteristic clinical and laboratory features. The routine evaluation of these lesions includes histologic and immunophenotypic analyses. In the small percentage of cases that pose diagnostic challenges, the algorithm can include molecular genetic studies to help render a diagnosis. Unique immunoglobulin heavy- and light-chain or T-cell-receptor gene rearrangements are used to establish clonality in B- and T-cell proliferations. Whereas clonality is usually a hallmark of a malignant process, the absence of clonality does not exclude malignancy. This is predicted on the disparate biology of the various lympho-proliferative processes and on the technical aspects of the tests used for their identification. As molecular genetic testing for gene rearrangements becomes commonplace in the clinical laboratory, the variety of methods for performing these tests has increased. Since molecular testing may be the deciding feature in making a diagnosis of malignancy, the purpose of this review is to discuss the advantages and problems of the polymerase chain reaction and Southern blot method in detecting immunoglobulin gene rearrangements in B cells. The added goal of this summary is to encourage the laboratorian to interpret these tests in the context of complete clinical and histologic data.