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Cellularity
Kinds of Cellularity Selected AbstractsPreparation of thyroid FNA material for routine cytology and BRAF testing: A validation studyDIAGNOSTIC CYTOPATHOLOGY, Issue 3 2010Giancarlo Troncone M.D., Ph.D. Abstract V600E BRAF mutation is emerging as an independent marker of papillary thyroid carcinoma aggressive behavior. Papillary thyroid carcinomas harboring this mutation should be extensively resected. However, this requires an unquestionable cytological diagnosis of malignancy. Thus, cytological specimens should be properly handled to provide both morphological and molecular information. Here, we assessed whether our method of preparation of fine-needle aspiration material is suitable for both tests. A series of 128, routinely performed, fine-needle aspirations was analyzed. Each nodule was punctured three times. A representative Diff-Quik smear prepared from the first two passages was evaluated onsite. When microscopy was diagnostic (n = 44), the third needle pass was dedicated to harvest material for BRAF testing; in the remaining cases (n = 84), additional direct smears for cytology were prepared and the remaining material in the needle plus the needle rinsing was collected for BRAF testing. Cellularity was adequate in 126/128 (98%) cases. Cytological diagnoses were inadequate (2%), benign (85%), follicular lesion of undetermined significance (5%), follicular neoplasms (2%), suspicious for malignancy (2%), and malignant (4%). Higher average of extracted DNA concentration was observed in the dedicated pass group (25.9 vs 7.95 ng/,l). However, the rate of successful exon 15 BRAF amplification was similar with (43/44; 97.7%) or without (79/84; 94%) the dedicated pass. Thus, our protocol is suitable for both tests. Whenever necessary BRAF testing may also be performed on the residual samples of thyroid nodules, without interfering with routine cytology. Diagn. Cytopathol. 2010. © 2009 Wiley-Liss, Inc. [source] Muscle Cellularity at Cranial and Caudal Levels of the Trunk Musculature of Commercial Size Sea Bass, Dicentrarchus labrax (Linnaeus, 1758)ANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 5 2005I. Abdel Summary In eight specimens of Atlantic sea bass of commercial size (,350 g) muscle cellularity was studied at two selected sampling levels of the trunk axial musculature: caudal (anal opening) and cranial (fourth radius of the dorsal fin). The following parameters were quantified at both sampling levels: white muscle cross-sectional area, white muscle fibre diameter (900,1200 fibres), muscle fibre number and muscle fibre density. Results showed a higher total cross-sectional area at cranial than at caudal level (P < 0.05), what is related with their different gross morphology. However, the white muscle fibre size distribution, as well as the muscle fibre number and density did not show significant differences between them. This study contributes to typify muscle fibre sampling in sea bass of commercial size what is of great interest for morphometric studies where white muscle cellularity is commonly correlated with textural or organoleptic parameters. [source] Effects of Liposome-Encapsulated Hemoglobin on Human Immune System: Evaluation in Immunodeficient Mice Reconstituted With Human Cord Blood Stem CellsARTIFICIAL ORGANS, Issue 2 2009Akira T. Kawaguchi Abstract As preclinical evaluation in animals does not necessarily portray human responses, liposome-encapsulated hemoglobin (LEH), an artificial oxygen carrier, was tested in immunodeficient mice reconstituted with human hematopoietic stem cells (cord blood-transfused NOD/SCID/IL-2R,null[CB-NOG] mice). Changes in immunocompetent T-cell and B-cell composition in peripheral blood, spleen, and bone marrow were examined 2 and 7 days after 10 mL/kg of intravenous administration of LEH, empty liposome (EL), or saline using immunohistochemical and flow cytometrical techniques in wild-type mice and CB-NOG mice. Responses to intraperitoneal administration of toxic shock syndrome toxin-1 (TSST-1) under the absence or presence of LEH (10 mL/kg) were also determined 4 h and 3 days later in terms of lymphocyte composition and IL-2 plasma level in wild-type as well as CB-NOG mice. When liposome (LEH or EL) was administered to wild-type or CB-NOG mice, the composition of B-cells and T-cells in the spleen or peripheral blood failed to show any consistent or significant changes. The responses to a bacterial antigen (TSST-1) measured by IL-2 production were comparable regardless of the presence or absence of LEH in wild-type as well as in CB-NOG mice. Cellularity, distribution, and maturation of these human cells in peripheral blood, spleen, and bone marrow were comparable among the groups. The results suggest that simple LEH administration may not change immune cellularity, and LEH presence may not largely affect the early T-cell response to bacterial enterotoxins in murine as well as in reconstituted human immune systems. [source] A CRITICAL LOOK AT PAP ADEQUECY: ARE OUR CRITERIA SATISFACTORY?CYTOPATHOLOGY, Issue 2006D.R. Bolick Liquid based Pap (LBP) specimen adequacy is a highly documented, yet poorly understood cornerstone of our GYN cytology practice. Each day, as cytology professionals, we make adequacy assessments and seldom wonder how the criteria we use were established. Are the criteria appropriate? Are they safe? What is the scientific data that support them? Were they clinically and statistically tested or refined to achieve optimal patient care? In this presentation, we will take a fresh look at what we know about Pap specimen adequacy and challenge some of the core assumptions of our daily practice. LBP tests have a consistent, well-defined surface area for screening, facilitating the quantitative estimates of slide cellularity. This provides an unprecedented opportunity to establish reproducible adequacy standards that can be subjected to scientific scrutiny and rigorous statistical analysis. Capitalizing on this opportunity, the TBS2001 took the landmark step to define specimen adequacy quantitatively, and set the threshold for a satisfactory LBP at greater than 5,000 well visualized squamous epithelial cells. To date, few published studies have attempted to evaluate the validity or receiver operator characteristics for this threshold, define an optimal threshold for clinical utility or assess risks of detection failure in ,satisfactory' but relatively hypocellular Pap specimens. Five years of cumulative adequacy and cellularity data of prospectively collected Pap samples from the author's laboratory will be presented, which will serve as a foundation for a discussion on ,Pap failure'. A relationship between cellularity and detection of HSIL will be presented. Risk levels for Pap failure will be presented for Pap samples of different cellularities. The effect of different cellularity criterion on unsatisfactory Pap rates and Pap failure rates will be demonstrated. Results from this data set raise serious questions as to the safety of current TBS2001 adequacy guidelines and suggest that the risk of Pap failure in specimens with 5,000 to 20 000 squamous cells on the slide is significantly higher than those assumed by the current criteria. TBS2001 designated all LBP to have the same adequacy criterion. Up to this point, it has been assumed that ThinPrep, SurePath, or any other LBP would be sufficiently similar that they should have the same adequacy criteria. Data for squamous cellularity and other performance characteristics of ThinPrep and SurePath from the author's laboratory will be compared. Intriguing data involving the recently approved MonoPrep Pap Test will be reviewed. MonoPrep clinical trial data show the unexpected finding of a strong correlation between abundance of endocervical component and the detection of high-grade lesions, provoking an inquiry of a potential new role for a quantitative assessment of the transition zone component. The current science of LBP adequacy criteria is underdeveloped and does not appear to be founded on statistically valid methods. This condition calls us forward as a body of practitioners and scientists to rigorously explore, clarify and define the fundamental nature of cytology adequacy. As we forge this emerging science, we will improve diagnostic performance, guide the development of future technologies, and better serve the patients who give us their trust. Reference:, Birdsong GG: Pap smear adequacy: Is our understanding satisfactory? Diagn Cytopathol. 2001 Feb; 24(2): 79,81. [source] Diagnostic pitfalls in the evaluation of fine needle aspiration cytology of the thyroid: correlation with histopathology in 260 casesCYTOPATHOLOGY, Issue 2 2009A. N. Haberal Objectives:, Fine needle aspiration cytology (FNAC) of the thyroid is a non-invasive, cost-effective screening procedure that is valuable for distinguishing neoplastic lesions from non-neoplastic nodules. The aim of this study was to determine the diagnostic accuracy of FNACs performed at our institution by correlating FNAC results with histopathological diagnoses. Methods:, Two hundred and seventy-one aspiration cytology specimens followed by thyroidectomy were included in the study, and the results of 260 adequate FNACs were compared with their histological diagnoses. Results:, The sensitivity and specificity of thyroid FNAC for detecting neoplasia were 92.6% and 91.6%, respectively. There were 15 (5.7%) false positives and six (2.3%) false negatives. Conclusions:, The results showed that follicular cells that exhibit some of the features of papillary carcinoma could be observed in a cytology slide of Hashimoto's thyroiditis, leading to a diagnostic pitfall. In addition, cellularity and overlapping cytological criteria in hyperplasia might lead to a false diagnosis. [source] A CRITICAL LOOK AT PAP ADEQUECY: ARE OUR CRITERIA SATISFACTORY?CYTOPATHOLOGY, Issue 2006D.R. Bolick Liquid based Pap (LBP) specimen adequacy is a highly documented, yet poorly understood cornerstone of our GYN cytology practice. Each day, as cytology professionals, we make adequacy assessments and seldom wonder how the criteria we use were established. Are the criteria appropriate? Are they safe? What is the scientific data that support them? Were they clinically and statistically tested or refined to achieve optimal patient care? In this presentation, we will take a fresh look at what we know about Pap specimen adequacy and challenge some of the core assumptions of our daily practice. LBP tests have a consistent, well-defined surface area for screening, facilitating the quantitative estimates of slide cellularity. This provides an unprecedented opportunity to establish reproducible adequacy standards that can be subjected to scientific scrutiny and rigorous statistical analysis. Capitalizing on this opportunity, the TBS2001 took the landmark step to define specimen adequacy quantitatively, and set the threshold for a satisfactory LBP at greater than 5,000 well visualized squamous epithelial cells. To date, few published studies have attempted to evaluate the validity or receiver operator characteristics for this threshold, define an optimal threshold for clinical utility or assess risks of detection failure in ,satisfactory' but relatively hypocellular Pap specimens. Five years of cumulative adequacy and cellularity data of prospectively collected Pap samples from the author's laboratory will be presented, which will serve as a foundation for a discussion on ,Pap failure'. A relationship between cellularity and detection of HSIL will be presented. Risk levels for Pap failure will be presented for Pap samples of different cellularities. The effect of different cellularity criterion on unsatisfactory Pap rates and Pap failure rates will be demonstrated. Results from this data set raise serious questions as to the safety of current TBS2001 adequacy guidelines and suggest that the risk of Pap failure in specimens with 5,000 to 20 000 squamous cells on the slide is significantly higher than those assumed by the current criteria. TBS2001 designated all LBP to have the same adequacy criterion. Up to this point, it has been assumed that ThinPrep, SurePath, or any other LBP would be sufficiently similar that they should have the same adequacy criteria. Data for squamous cellularity and other performance characteristics of ThinPrep and SurePath from the author's laboratory will be compared. Intriguing data involving the recently approved MonoPrep Pap Test will be reviewed. MonoPrep clinical trial data show the unexpected finding of a strong correlation between abundance of endocervical component and the detection of high-grade lesions, provoking an inquiry of a potential new role for a quantitative assessment of the transition zone component. The current science of LBP adequacy criteria is underdeveloped and does not appear to be founded on statistically valid methods. This condition calls us forward as a body of practitioners and scientists to rigorously explore, clarify and define the fundamental nature of cytology adequacy. As we forge this emerging science, we will improve diagnostic performance, guide the development of future technologies, and better serve the patients who give us their trust. Reference:, Birdsong GG: Pap smear adequacy: Is our understanding satisfactory? Diagn Cytopathol. 2001 Feb; 24(2): 79,81. [source] Fine needle aspiration cytology of follicular variant of papillary carcinoma of thyroidCYTOPATHOLOGY, Issue 4 2003M. Powari In this retrospective study, we tried to ascertain the fine needle aspiration cytology (FNAC) features of six histopathologically proven cases of the follicular variant of papillary carcinoma of thyroid (FVPCT). These proven cases were diagnosed from 1998,2000. May,Grunwald,Giemsa and haematoxylin & eosin stained FNAC smears were studied independently by two observers (MP and PD) for detailed cytological features. A comparison of the cytological features was undertaken with those reported in the literature. There were six cases of which only one case was diagnosed as FVPCT while the other five cases were diagnosed as follicular neoplasm (four cases) and neoplasm unclassifiable (one case) on FNAC smears. All these cases showed abundant cellularity with a prominent follicular pattern. No papillae were identified in any of the cases. Syncytial clusters (five cases), nuclear grooves (six cases), nuclear inclusions (one case) and chewing gum colloid (three cases) were noted in variable proportions. We suggest that a differential diagnosis of FVPCT should be considered if the cytology smears show abundant cellularity, syncytial clusters and follicular arrangement along with thick colloid. [source] Peripheral endothelial cells are not reliable in differentiating primary benign and malignant hepatocellular lesions in fine needle aspirates of the liverCYTOPATHOLOGY, Issue 3 2002GORDON H. YU The distinction of hepatocellular carcinoma (HCC) from benign lesions of the liver in fine needle aspiration (FNA) specimens can be problematic. In an attempt to separate well-differentiated HCC from benign hepatocellular lesions, the presence of tissue fragments displaying peripheral endothelial cells (PE) has been proposed in a previous study as a useful feature in favour of malignancy. In this study, we evaluated slides from 59 cases of liver masses undergoing FNA (19 HCC, 40 benign) and evaluated them for the presence of tissue fragments containing PE. We found that 90% of cases of HCC contained tissue fragments in which PE were either focally present or abundant. However, 68% of cases containing only benign hepatocytes also contained tissue fragments in which PE were at least focally present. In addition, it appears that within the group of benign lesions, the presence of PE was related to the overall cellularity of the specimen rather than the specific nature of the lesion. Thus, the presence of PE in tissue fragments does not, in isolation, appear to be a useful morphological feature for the separation of benign and malignant hepatocellular lesions in FNA material. [source] Does Imiquimod Histologically Rejuvenate Ultraviolet Radiation,Damaged Skin?DERMATOLOGIC SURGERY, Issue 12 2007KATHLEEN SMITH MD BACKGROUND Imiquimod (IMI) 5% is believed by some to result in an improved cosmetic appearance of chronically ultraviolet radiation (UV)-damaged skin. OBJECTIVE The objective was to determine what histologic and immunohistologic changes were present in actinically damaged skin after treatment with IMI. METHODS AND MATERIALS Pre- and posttherapy biopsies of 12 patients with histories of actinic keratoses were evaluated with routine histology and immunohistochemical stains including p53, p63, proliferating cell nuclear antigen (PCNA), c-kit, and Factor XIIIa. RESULTS After IMI therapy there was less compact hyperkeratosis, a more uniform rete ridge pattern with a more ordered proliferation of the epidermis, and a decrease in sun-damaged melanocytes. The papillary dermis showed a more uniform cellularity, and there was increased cellularity within the area of solar elastosis. After therapy, staining for p53, p63, and PCNA was decreased within the epidermis; staining for c-kit was decreased but more uniform in the basal cell; and Factor XIIIa expression was increased within the papillary dermis with a more ordered pattern of staining. CONCLUSION These morphologic and immunohistochemical patterns may explain some of the improvement in overall skin appearance after IMI therapy and may be related to the spectrum of signaling pathways induced by the imidazoquinolines. [source] Conjunctival impression cytology in trachomaDIAGNOSTIC CYTOPATHOLOGY, Issue 3 2009Anshu M.D., D.N.B. Abstract Trachoma is one of the leading causes of blindness and clinical examination remains the mainstay of diagnosis. However there is need to evaluate simple, inexpensive techniques which can be used for screening of trachoma in endemic regions. We report two cases where conjunctival impression cytology played a part in confirming the diagnosis of trachoma. We used a modified technique of obtaining conjunctival impressions, which not only met with better patient compliance and minimal ocular distress, but also provided better cellularity and morphology of cells for evaluation. The impression smears showed squamous metaplasia and loss of goblet cells. The cytoplasm of these cells had a hazy, moth eaten appearance and showed presence of intracytoplasmic inclusions. These basophilic inclusions were present singly and in clusters and were around 5 ,m in diameter. A large number of these inclusions were present extracellularly as well. Conjunctival impression cytology is a simple procedure which needs to be evaluated for its potential to be used for screening trachoma in endemic areas. Diagn. Cytopathol. 2009. © 2009 Wiley-Liss, Inc. [source] Epidermal inclusion cyst: Cytomorphological features and differential diagnosisDIAGNOSTIC CYTOPATHOLOGY, Issue 12 2008Uma Handa M.D. Abstract Aspirates from 162 epidermal inclusion cysts (EIC) from 157 patients were analyzed in order to elaborate on specific cytologic features. The most common site involved was the head and neck region (96 cases; 59.2%). The maximum patients were in the 3rd and 4th decades of life. Aspirates from EIC showed a clear background, with high cellularity, and nucleate and anucleate squames. Keratinous material was seen in some cases but the amount was less compared with the cellular elements. In 31 cases, a diagnosis of infected EIC was made on the basis of dense inflammatory infiltrate in addition to the squames. Histopatholgy was available in 56 cases out of which EIC was diagnosed in 45 cases. The remaining 11 cases were dermoid cyst (5 cases), branchial cyst (2 cases), pilomatricoma (2 cases), and sebaceous and thyroglossal cyst (1 case each). Thus, EIC should be differentiated from other squamous cell containing lesions. Diagn. Cytopathol. 2008. © 2008 Wiley-Liss, Inc. [source] Cytologic feature by squash preparation of pineal parenchyma tumor of intermediate differentiationDIAGNOSTIC CYTOPATHOLOGY, Issue 10 2008Keiji Shimada M.D., Ph.D. Abstract Pineal parenchyma tumor of intermediate differentiation (PPTID) is a very rare intracranial tumor, and pathological investigation limited to immunohistological and ultrastructural analyses have been published to date. Although intraoperative cytology is one of the important approaches for initial diagnosis in brain tumors, no or little studies on cellular morphology of PPTID have been demonstrated due to its rarity. We report here cytological features of PPTID obtained from stereotactic surgical specimens in a case of 27-year-old female manifested by dizziness and diplopia. Brain MRI revealed an unhomogeneously enhanced, large-sized tumor (56 × 52 × 60 mm) mainly located in the pineal region expanding from the midbrain to superior portion of the cerebellum and the fourth ventricle. Squash cytology showed increased nucleocytoplasmic ratio, hyperchromatic nuclei, and small rosette-like cell cluster but cellular pleomorphism was mild to moderate and necrotic background was not observed. Histology showed high cellularity, moderate nuclear atypia, and small rosette formation but neither bizarre tumor cells nor necrosis was present. Mitotic counts were very low (less than 1 per 10 high-power fields) and the MIB-1 labeling index was relatively high (10.1%). Tumor cells were immunohistochemically positive for neural markers such as synaptophysin, neurospecific enolase but not for glial fibrillary acidic protein or S-100. In some parts, cells were strongly reactive for neurofilament protein. Taken together, we made a final diagnosis of PPTID. This is the first presentation of cytological analysis by squash preparation that gives an important clue to accurate diagnosis of pineal parenchymal tumor and to understand its malignant potential. Diagn. Cytopathol. 2008;36:749,753. © 2008 Wiley-Liss, Inc. [source] Do we need more than one ThinPrep to obtain adequate cellularity in fine needle aspiration?DIAGNOSTIC CYTOPATHOLOGY, Issue 11 2007Farnaz Hasteh M.D. First page of article [source] Comparison of cell block preparation methods for nongynecologic ThinPrep specimensDIAGNOSTIC CYTOPATHOLOGY, Issue 10 2007Kelly Nigro M.D. Abstract The purpose of this study was to compare four cell block (CB) methods in the setting of nongynecologic ThinPrep (TP) specimens. 48 CBs were prepared from 12 nongynecologic TP specimens using the following CB methods: (1) Inverted filter sedimentation (IFS); (2) Thrombin method; (3) Albumin method; (4) Simple sedimentation. Each CB was assigned a cellularity score: 0 no cells, 1+ hypocellular, 2+ hypocellular with tissue fragments, 3+ cellular. A score of 2+ or 3+ was given for 11/12 of thrombin, 7/12 IFS, 5/12 albumin, and 2/12 simple sedimentation CBs. Thrombin CBs demonstrated a pale background clot with evenly distributed cells. Albumin CBs had a cracked uneven background. IFS CBs had a clear background, but were technically difficult and cells appeared artifactually crowded. In the setting of nongynecologic TP specimens, the thrombin CB was easily prepared and produced the best CB in regards to cellularity, cell distribution, and background quality. Diagn. Cytopathol. 2007;35:640,643. © 2007 Wiley-Liss, Inc. [source] Utility of cell blocks in the diagnosis of thyroid aspiratesDIAGNOSTIC CYTOPATHOLOGY, Issue 2 2006Niria Sanchez M.D. Abstract Cell blocks (CBs) are often prepared with fine-needle aspirates (FNAs) from multiple organs as an adjunct to smears in the diagnosis of aspirated lesions. However, the literature contains few reports on their utility with regard to specific organ sites. At our institution, CBs are made routinely on FNAs when there is sufficient material remaining after smear preparation, with thyroid representing the largest volume. The aim of this study was to determine the utility of CBs in the diagnosis of thyroid lesions. From January 2002 to April 2004, 546 thyroid FNAs were performed. Eighty-two (15%) cases, from 60 females and 20 males (age range, 17,88 yr; mean, 50 yr), had CBs and formed the basis of this study. Seventy-four (90%) of the cases were performed by the radiologist or the clinician and 8 (10%) by the pathologist, all of which had an immediate assessment for adequacy. One to 7 passes were performed with an average of 3/case. The needles were immediately rinsed in Hanks' Balanced Salt Solution after smear preparation. CBs were made on bloody specimens/those with tissue fragments. Cell-block slides were reviewed for the presence of cellular elements and classified into three categories: (1) contributory, (2) noncontributory, or (3) provides additional information. Of the 82 cases, 23 (28%) were neoplastic, 51 (62%) were nonneoplastic, and 8 (10%) were nondiagnostic. Fifteen of the neoplastic cases had confirmatory biopsies, 9 of which were papillary carcinoma. The overall cellularity of the CBs was low, varying from 0 to 2 follicular groups in the noncontributory CBs and 3 to 6 follicular groups or papillary formations in the contributory CBs. CBs were contributory in 25 (31%) cases: 5 neoplastic (1 follicular neoplasm, 3 papillary carcinoma, and 1 suspicious for papillary carcinoma), 18 nonneoplastic, and 2 nondiagnostic. CBs were noncontributory in 56 (68%) cases: 18 neoplastic (4 papillary carcinomas, 1 suspicious for papillary carcinoma, 4 Hürthle cell neoplasms, and 9 follicular neoplasms), 33 nonneoplastic, and 5 nondiagnostic. One case was categorized as provided additional information because the CB showed material that was not present on the slides; however, it was still nondiagnostic. In summary, CBs did not help in the majority of cases. They were contributory in only 25 (31%) of the 82 cases, and of the 23 neoplastic cases, only 5 (22%) CBs were contributory. The contribution of the CBs in the diagnosis of thyroid lesions was minimal because of the low cellularity. On-site assessment of specimen adequacy often results in fewer passes, thus contributing to the low cellularity present in cell-block preparations. Ancillary studies may require additional passes. Diagn. Cytopathol. 2006; 34:89,92. © 2006 Wiley-Liss, Inc. [source] The significance of the diagnosis of atypia in breast fine-needle aspirationDIAGNOSTIC CYTOPATHOLOGY, Issue 5 2004Jennifer C. Lim M.D. Abstract The diagnosis of atypia in breast fine-needle aspiration (FNA) continues to be an area of debate in cytology practice. The aim of this study was to assess the clinical significance of this term and to evaluate potential morphological criteria, which would determine the patient's outcome. A computer-based search was carried out to retrieve breast FNAs performed between 1990 and 2000 that were diagnosed as atypical. Cases followed by surgical resection were reexamined for the presence of morphological features potentially differentiating benign and malignant lesions. Out of 1,568 breast FNAs, there were 64 cases (4%) with a diagnosis of atypia. Thirty-eight cases had surgical follow-up material that revealed malignancy in 14 cases (37%) and benign lesions in 24 cases (63%). The benign diagnostic categories included fibrocystic change (12/24), fibroadenoma (3/24), tubular adenoma (2/24), and nonspecific findings (7/24). The malignant diagnoses included ductal carcinoma (9/14), lobular carcinoma (3/14), ductal carcinoma in situ (DCIS; 1/14), and tubular carcinoma (1/14). The evaluation of cytological criteria used to differentiate benign from malignant lesions (i.e., cellularity, loss of cohesion, myoepithelial cells, nuclear enlargement, nuclear overlap, prominent nucleoli) revealed significant overlap between benign and malignant cases, particularly in cases of fibroadenoma, tubular adenoma, and proliferative breast disease. The surgical follow-up of four hypocellular cases revealed lobular carcinoma in two cases and ductal carcinoma in the remaining two cases. Our study confirmed that the diagnosis of atypia is clinically significant because it is associated with a high probability of malignancy. No morphological criterion is able to reliably differentiate benign and malignant lesions in cases diagnosed with atypia. Diagnosis of atypia is particularly significant in hypocellular cases. We recommended that breast FNAs with a diagnosis of atypia be evaluated further histologically. Diagn. Cytopathol. 2004;31:285,288. © 2004 Wiley-Liss, Inc. [source] Low-grade urothelial carcinoma: Reappraisal of the cytologic criteria on ThinPrep®DIAGNOSTIC CYTOPATHOLOGY, Issue 3 2003Ph.D., Wei Xin M.D. Abstract The diagnostic criteria for low-grade urothelial lesions that have been described in the past were based on urinary specimens prepared by the cytospin method. Recognizing the recent popularity of the ThinPrep® methodology and the cytologic alterations it introduces to the cellular features, we sought to evaluate the reproducibility of these criteria in ThinPrep urinary samples. One hundred twenty-six ThinPrep urinary specimens with a tissue diagnosis of low-grade urothelial carcinoma (LGUC) and 45 negative controls were evaluated. Three pathologists blindly reviewed the slides separately and the consensus on each feature was used in the study. Logistic regression analysis was used to determine which criteria in combination were most predictive of low-grade urothelial carcinoma. All specimens were evaluated for the following 18 features: nucleus/cytoplasm ratio, irregular nuclear border, cytoplasm homogeneity, cell clusters, high cellularity, prominent nucleoli, granular nuclear chromatin, hyperchromasia, acute inflammation, vesicular chromatin, nuclear molding, nuclear eccentricity, elongated nuclei, necrosis, anisonucleosis, irregular bordered fragments, absent cytoplasmic collar, and peripheral palisading. High nucleus-to-cytoplasm ratio, irregular nuclear borders, and homogeneous cytoplasm (combination sensitivity of 59% and specificity of 100%) were the best predictive features for LGUC. Minor predictive criteria were eccentric nuclei and nuclear molding. ThinPrep provides well preserved, cleaner specimens without significantly altering the morphology. The three key criteria applied in cytospin specimens to diagnose LGUC were reproducible in ThinPrep specimens. Diagn. Cytopathol. 2003;29:125,129. © 2003 Wiley-Liss, Inc. [source] Utility of cytomorphologic criteria and p53 immunolocalization in distinguishing benign from malignant cystic squamous-lined lesions of the neck on fine-needle aspirationDIAGNOSTIC CYTOPATHOLOGY, Issue 1 2002Joseph F. Nasuti M.D. Abstract Fine-needle aspiration can effectively distinguish between benign and malignant cystic lesions of the head and neck. However, in some instances it may be difficult to arrive at a definite diagnosis due to limited cellularity, reactive changes, and cellular degeneration. In this study we examined the usefulness of six cytomorphologic features including the presence or prevalence of nuclear atypia, anucleated cells, tissue fragments, necrosis, and background inflammation in distinguishing between benign and malignant cystic lesions of the head and neck. The case cohort comprised 14 benign and 22 malignant cases. P53 immunostain was performed in 19 cases. These features were semiquantitatively measured on a sliding scale of 0,4 in both air-dried Diff-Quik-stained, alcohol-fixed Papanicolaou-stained smears and Millipore filter preparations. Mean and standard errors were calculated and statistical significance was evaluated by unpaired t -test (StatView). Increased number of tissue fragments (P < 0.001), greater degree of nuclear atypia (P < 0.001), and background necrosis (P < 0.001) were more frequent in cystically degenerating squamous carcinoma as compared to benign squamous cystic lesions. No significant differences were noted in the number of single cells, anucleated cells, or in the amount of background inflammation found in aspirates of benign vs. malignant cystic squamous lesions. A higher percentage of the malignant cystic squamous lesions FNA cases demonstrated p53 immunolocalization but this difference was not statistically significant. Application of the above-mentioned cytomorphologic criteria and the use of p53 immunostain could effectively distinguish between benign and malignant cystic lesions of the head and neck. Diagn. Cytopathol. 2002;27:10,14. © 2002 Wiley-Liss, Inc. [source] Diagnosis of melanoma aspirates on ThinPrep®: The University of Michigan experienceDIAGNOSTIC CYTOPATHOLOGY, Issue 5 2002Güliz Akdas Barkan M.D. Abstract The purpose of this study was to compare the cytologic features of melanoma fine-needle aspirates (FNAs) prepared by ThinPrep® (TP) with those in conventional smears (CS) and to identify any diagnostic pitfalls. Fifty-one aspirates diagnosed as melanoma were obtained, 36 of which were prepared by both TP and CS. The preparations were evaluated for cellularity, cell aggregates, cellular appearance, melanin pigment, cytoplasmic, and nuclear features. Categorical data were analyzed by the chi-square test and continuous data by the Wilcoxin-signed rank test. Correlation was determined by Spearman's test for bivariate correlations (rho). Good correlation between the two methods was identified for the following features: cellularity, cell type, bi/multinucleated cells, cytoplasmic features, NC ratio, and presence of macronucleoli. TP exhibits coarser chromatin compared to CS (P = 0.005). Six of 36 CS contained large cellular groups; none of the TP contained them (P = 0.018). Twenty-five of 36 CS contained intranuclear inclusions as opposed to 12/36 TP (P < 0.001). The number of inclusions was significantly reduced on TP. The amount of intracellular melanin was the same with both techniques. Background melanin was markedly reduced on TP except when either trapped by fibrin or attached to cellular clusters (P = 0.006). Background blood was also markedly reduced on TP (P < 0.005). In summary, the cytological features of TP and CS for FNA evaluation of melanoma correlate well; however, one needs to be aware of the cytologic alterations introduced by TP. TP is a sufficient preparation method in the diagnosis of melanoma FNA aspirates when performed by clinicians. It is also a useful adjunct in bloody or low-cellular aspirates, where it tends to reduce the background blood and concentrate the cells. Diagn. Cytopathol. 2002;26:334,339. © 2002 Wiley-Liss, Inc. [source] Breast masses in males: Multi-institutional experience on fine-needle aspirationDIAGNOSTIC CYTOPATHOLOGY, Issue 2 2002Momin T. Siddiqui M.D. Abstract Male breast masses are uncommon pathologic findings. They are rarely aspirated, resulting in limited cytopathologic experience. The following study describes the cytopathology of male breast lesions from data collected for a period of 10 yr from three large institutions. A total of 14,026 breast aspirations were performed of which 614 were from male patients. All cases were reviewed and correlated with the appropriate clinicopathologic follow-up. The FNA diagnoses were as follows: benign, 427 cases (gynecomastia 353, fat necrosis 21, miscellaneous 53); malignant, 32 cases (ductal carcinoma nos 15, metastatic tumors 17); and atypical/suspicious, 61 cases. Ninety-four cases were nondiagnostic due to scant cellularity. Male breast aspirates accounted for 4.3% of the total breast FNAs performed. The clinicopathologic follow-up in both the benign and malignant categories showed 100% correlation. The overall sensitivity was 95.3%, specificity was 100%, and diagnostic accuracy was 98%. A relatively high specimen unsatisfactory rate was seen (>15%). The commonest cytopathologic diagnosis was gynecomastia, followed by ductal carcinoma. Florid duct atypia in gynecomastia may mimic adenocarcinoma, necessitating a higher threshold for cytopathologic interpretation for malignancy in males. Diagn. Cytopathol. 2002;26:87,91; DOI 10.1002/dc.10066 © 2002 Wiley-Liss, Inc. [source] Morphologic predictors of papillary carcinoma on fine-needle aspiration of thyroid with ThinPrep® preparationsDIAGNOSTIC CYTOPATHOLOGY, Issue 6 2001Yilin Zhang M.D. Abstract Although the cytologic features of papillary carcinoma of the thyroid are well-known, none is entirely specific. We conducted this study to determine the minimal criteria necessary to achieve 100% specificity for the diagnosis of papillary carcinoma on fine-needle aspiration (FNA). Forty patients with histologically confirmed papillary carcinoma and 17 patients with other thyroid lesions who underwent preoperative FNA at Beth Israel Deaconess Medical Center during a 4-yr period were included in the study. All cytology slides were prepared with the ThinPrep® processing technique. Various architectural and nuclear features were evaluated, with a score assigned to each feature, and correlated with the histologic diagnosis of papillary carcinoma. Intranuclear inclusions, papillary and/or sheet arrangements, nuclear grooves, powdery chromatin, nuclear molding, high cellularity, and small nucleoli were significantly associated with papillary carcinoma (P < 0.05). The requirement of any intranuclear inclusions and many nuclear grooves, or a minimum of sum of scores (of the above eight features) of 10, yields 100% specificity and approximately 70% sensitivity. Cases with fewer features can be reported as suspicious or indeterminate for papillary carcinoma. A quantitative/probabilistic approach in the reporting of thyroid FNA provides a practical guide for management of patients with thyroid nodules. Diagn. Cytopathol. 24:378,383, 2001. © 2001 Wiley-Liss, Inc. [source] Prognostic factors in advanced stage Hodgkin's lymphoma: the significance of the number of involved anatomic sitesEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 5-6 2001T.P. Vassilakopoulos Abstract:Background: Advanced Hodgkin's lymphoma (HL) is curable by conventional chemotherapy in 60,70% of patients. The pretreatment identification of a sizeable subgroup of patients with sufficiently low failure-free survival (FFS) to be eligible for investigational treatment is necessary. Objectives: To determine the prognostic significance of the number of involved sites (NIS) in patients with advanced HL and its relationship to the International Prognostic Score (IPS). Methods: A retrospective review of patients with advanced HL, defined as Ann Arbor stage (AAS) IB, IIB, III or IV, treated with anthracycline-based regimens. The end-point was FFS. Results: We identified 277 patients with a median age of 32 yr (14,78), 57% of whom were males. AAS was I in 4% of patients, II in 29%, III in 38% and IV in 29%. B-symptoms were recorded in 81%. Most patients had nodular sclerosis (64%) and mixed cellularity (26%) histology. IPS was ,3 in 44% of 242 evaluable patients. The NIS was ,5 in 32% of the patients and 20% of all patients had both ,5 involved sites and IPS ,3. The 10-yr FFS was 67%, being 76% vs. 50% for patients with ,4 vs. ,5 involved sites (P < 0.0001). The NIS (, 5), AAS IV and anemia were independent predictors of FFS in multivariate analysis. The NIS remained significant along with IPS, when the latter was included in the analysis. Patients with ,5 involved sites and IPS ,3 had 10-yr FFS overall, and relapse-free survival of 41%, 45% and 49%, respectively. Conclusions: The NIS was associated with FFS in advanced HL, was independent of IPS, and led to the identification of a sizeable subgroup of patients with 10-yr FFS of approximately 40%. This factor should be evaluated during the development of prognostic systems. [source] CREB function is required for normal thymic cellularity and post-irradiation recoveryEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 7 2004Sven Baumann Abstract Recent generation of genetically modified Creb1 mutant mice has revealed an important role for CREB (cAMP responsive element binding protein) and the related proteins CREM (cAMP responsive element modulator) and ATF1 (activating transcription factor 1) in cell survival, in agreement with previous studies using overexpression of dominant-negative CREB (dnCREB). CREB and ATF1 are abundantly expressed in T cells and are rapidly activated by phosphorylation when T cells are stimulated through the T cell antigen receptor. We show that T cell-specific loss of CREB in mice, in combination with the loss of ATF1, results in reduced thymic cellularity and delayed thymic recovery following sublethal irradiation but no changes in T cell development or activation. These data show that loss of CREB function has specific effects on thymic T lymphocyte proliferation and homeostasis in vivo. [source] Impaired lymphocyte development and function in Clast5/Stra13/DEC1-transgenic miceEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 5 2004Mika Seimiya Abstract Clast5/Stra13/DEC1 is a member of the helix-loop-helix family of transcriptional repressors. We have previously shown that Clast5 is rapidly down-regulated upon B,cell activation and its overexpression inhibits cell cycle progression in B,lymphoma cells. In the present study, we show that Clast5 expression is developmentally regulated during B,cell differentiation, being expressed at theprogenitor B,cells, down-regulated at the precursor B,cells, elevated in immature and mature resting B,lymphocytes, and down-regulated again in germinal center B,ells. To investigate the function of Clast5 in regulating lymphocyte development, we have generated transgenic mice expressing Clast5 in B- and T-lineage cells (Clast5-Tg). Clast5-Tg mice grew and bred normally but their spleen and thymus cellularity was reduced compared with control littermates. The development of B,cells in the bone marrow and T,cells in the thymus was impaired, with the expansion of progenitor B and T,cells most strongly affected. The frequency of IL-7-responsive cells in the bone marrow of Clast5-Tg mice was reduced by >80% and their proliferative response to IL-7 was also compromised. Mature B,cells from Clast5-Tg mice were hyporesponsive to antigen receptor cross-linking and exhibited mild reduction in the proliferative response to CD40 ligation or lipopolysaccharide stimulation. Moreover, thedevelopment of germinal center B,cells and antibody production against a T-dependent antigen were reduced in Clast5-Tg mice. These results reveal a critical role for Clast5/Stra13/DEC1 in negatively regulating lymphocyte development and function in vivo. [source] Astrocyte-specific expression of a soluble form of the murine complement control protein Crry confers demyelination protection in the cuprizone modelGLIA, Issue 14 2007Dustin T. Briggs Abstract Complement has been implicated as a potential effector mechanism in neurodegeneration; yet the precise role of complement in this process remains elusive. In this report, we have utilized the cuprizone model of demyelination-remyelination to examine the contribution of complement to disease. C1q deposition was observed in the corpus callosum of C57BL/6 mice during demyelination, suggesting complement activation by apoptotic oligodendrocyte debris. Simultaneously, these mice lost expression of the rodent complement regulatory protein, Crry. A soluble CNS-specific form of the Crry protein (sCrry) expressed in a transgenic mouse under the control of an astrocyte-specific promoter was induced in the corpus callosum during cuprizone treatment. Expression of this protein completely protected the mice from demyelination. Interestingly, sCrry mice had low levels of demyelination at later times when control mice were remyelinating. Although the sCrry transgenic mice had lower levels of demyelination, there was no decrease in overall cellularity, however there were decreased numbers of microglia in the sCrry mice relative to controls. Strikingly, sCrry mice had early recovery of mature oligodendrocytes, although they later disappeared. TUNEL staining suggested that production of the sCrry protein in the transgenic mice protected from a late apoptosis event at 3 weeks of cuprizone treatment. Our data suggest complement provides some protection of mature oligodendrocytes during cuprizone treatment but may be critical for subsequent remyelination events. These data suggest that temporal restriction of complement inhibition may be required in some disease settings. © 2007 Wiley-Liss, Inc. [source] Quantitative determination of the diagnostic accuracy of the synovitis score and its componentsHISTOPATHOLOGY, Issue 3 2010Elisabeth Slansky Slansky E, Li J, Häupl T, Morawietz L, Krenn V & Pessler F (2010) Histopathology,57, 436,443 Quantitative determination of the diagnostic accuracy of the synovitis score and its components Aims:, To assess the diagnostic accuracy of a three-component synovitis score and to determine the relative contribution of each of its components to its overall discriminatory power. Methods and results:, The synovitis score was determined in 666 synovial specimens: normal synovium, n = 33; post-traumatic arthropathy (PtA), n = 29; osteoarthritis (OA), n = 221; psoriatic arthritis (PsA), n = 42; and rheumatoid arthritis (RA), n = 341. The discriminatory abilities of the score and its components were quantified with binary and multicategory receiver operating characteristic (ROC) analysis. The score differentiated all arthropathies accurately from normal tissue (area under the ROC curve, AUC: 0.87,0.98) and RA from OA or PtA (AUC: 0.85 for both), but could not distinguish well within pairs of inflammatory or degenerative arthropathies. AUCs of the intimal hyperplasia and stromal cellularity components correlated with the AUCs of the complete score markedly more strongly (r = 0.94 and 0.91, respectively) than the inflammatory infiltration component (r = 0.60). Multicategory ROC analysis ranked the score several-fold higher than any of its components, and the components in the order stromal cellularity>intimal hyperplasia>infiltration. Conclusion:, Combining three distinct histological parameters into a three-component score produces greatly increased overall diagnostic power. The discriminatory ability of the score stems more from measuring proliferative than infiltrative aspects of synovitis. [source] Immunocytochemistry in liquid-based cervical cytology: Analysis of clinical use following a cross-sectional studyINTERNATIONAL JOURNAL OF CANCER, Issue 5 2006Shaira Sahebali Abstract Cytological screening for cervical cancer is hampered by imperfect sensitivity and low inter-observer reproducibility. Human papillomavirus (HPV) testing lacks specificity as a primary screening method. Studies indicate that immunocytochemical detection of alterations caused by HPV in the host cells can optimise screening. Here, the potential of p16INK4a (cyclin-dependent kinase inhibitor p16) and MIB-1 (Ki-67 proliferation marker) as adjunct molecular markers for cervical lesions was investigated in a prospective, cross-sectional study of 500 samples in the framework of opportunistic screening in Flanders, Belgium. A consecutive series of 200 samples and 100 samples from the cytological categories ASC, LSIL and HSIL were investigated. Surepath samples were interpreted according to the Bethesda 2001 reporting system. HPV testing was done with MY09/MY11 consensus PCR. Immunocytochemistry for p16INK4a and MIB-1 was performed with an automated staining protocol. The number of immunoreactive cells/1,000 cervical cells was assessed. There was a higher mean number of p16INK4A and MIB-1 immunoreactive cells/1,000 cells in HSIL (4.06 ± 1.93 and 11.13 ± 2.83, respectively) compared to other cytological categories. Both markers showed a large spread in counts, for all categories. In cases of HSIL without immunoreactive cells for either marker, low cellularity and long-term storage in water were often the cause of false negativity. This study confirms that positive staining for p16INK4a and MIB-1 is highly correlated with presence of high-grade lesions. These markers could be used as adjuncts to increase the sensitivity of cytological screening as well as the specificity of the HPV test. However, clear methodological standards are needed for optimal performance of immunocytochemistry in a clinical setting. © 2005 Wiley-Liss, Inc. [source] Metastatic cutaneous leiomyosarcoma from primary neoplasm of the mesenteryINTERNATIONAL JOURNAL OF DERMATOLOGY, Issue 8 2001Kyoung Jin Kim MD A 31-year-old South Korean woman was referred to the dermatology department from the oncology department for the evaluation of a subcutaneous nodular lesion on the back. Three years before, she noted a palpable, fingertip-sized, nontender mass on her right lower abdomen. The mass had increased in size slowly. One year ago, she visited a local clinic and physical examination revealed a 7 × 8 × 7 cm, slightly tender, deep-seated mass on the right lower quadrant of the abdomen. The mass on the ilial mesentery was resected by surgical exploration and tissue examination revealed leiomyosarcoma. She refused adjuvant chemotherapy. Approximately 3 months later, she re-visited the clinic with a tender, subcutaneous nodule on the back. Cutaneous examination revealed a solitary, 2 × 2 cm, well-defined, hard, movable, subcutaneous nodule on the upper back without skin color change (Fig. 1). She complained of tenderness on touching the lesion. Histologic examination of a biopsy specimen showed irregularly arranged spindle cells scattered throughout the dermis. They were arranged in haphazardly oriented or interweaving fascicles. Most of the spindle cells possessed elongated nuclei with blunt ends and some cells had a polygonal outline with irregularly shaped nuclei (Fig. 2). There were many mitoses: 3,4 per high-power (× 400) field. Immunohistochemically, smooth muscle actin and desmin were positive in most of the tumor cells (Fig. 3). S-100 reactivity was not observed. A diagnosis of metastatic leiomyosarcoma was made. About 1 month later, computed tomography showed two, ill-defined, heterogeneous, low attenuation masses in the right lobe of the liver, suggesting liver metastasis. The patient was treated with chemotherapy for 2 months and remains in good condition. Figure 1. 2 × 2 cm, solitary, well-defined, hard, movable, subcutaneous nodule without any overlying skin change Figure 2. (a) Characteristic findings of cutaneous leiomyosarcoma with markedly high cellularity and densely packed transverse and longitudinal fascicles of cells (hematoxylin and eosin, × 40). (b) High magnification of the neoplasm revealing spindle cells with blunt-ended nuclei, pleomorphism, and mitotic figures (hematoxylin and eosin, × 200) Figure 3. Dense cytoplasmic reactivity for smooth muscle actin is apparent (smooth muscle actin, × 200) [source] Popliteal lymph node assay: facts and perspectivesJOURNAL OF APPLIED TOXICOLOGY, Issue 6 2005Guillaume Ravel Abstract The popliteal lymph node assay (PLNA) derives from the hypothesis that some supposedly immunemediated adverse effects induced by certain pharmaceuticals involve a mechanism resembling a graft-versus-host reaction. The injection of many but not all of these compounds into the footpad of mice or rats produces an increase in the weight and/or cellularity of the popliteal lymph node in the treated limb (direct PLNA). Some of the compounds known to cause these adverse effects in humans, however, failed to induce a positive PLNA response, leading to refinements of the technique to include pretreatment with enzyme inducers, depletion of CD4+ T cells or additional endpoints such as histological examination, lymphocyte subset analysis and cytokine fingerprinting. Alternative approaches have been used to improve further the predictability of the assay. In the secondary PLNA, the test compound is injected twice in order to illicit a greater secondary response, thus suggesting a memory-specific T cell response. In the adoptive PLNA, popliteal lymph node cells from treated mice are injected into the footpad of naive mice; a marked response to a subsequent footpad challenge demonstrates the involvement of T cells. Finally, the reporter antigens TNP-Ficoll and TNP-ovalbumin are used to differentiate compounds that induce responses involving neo-antigen help or co-stimulatory signals (modified PLNA). The PLNA is increasingly considered as a tool for detection of the potential to induce both sensitization and autoimmune reactions. A major current limitation is validation. A small inter-laboratory validation study of the direct PLNA found consistent results. No such study has been performed using an alternative protocol. Other issues include selection of the optimal protocol for an improved prediction of sensitization vs autoimmunity, and the elimination of false-positive responses due to primary irritation. Finally, a better understanding of underlying mechanisms is essential to determine the most relevant endpoints. The confusion resulting from use of the PLNA to predict autoimmune-like reactions as well as sensitization should be clarified. Interestingly, most drugs that were positive in the direct PLNA are also known to cause drug hypersensitivity syndrome in treated patients. This observation is expected to open new avenues of research. Copyright © 2005 John Wiley & Sons, Ltd. [source] Methods for the identi,cation of chemical respiratory allergens in rodents: comparisons of cytokine pro,ling with induced changes in serum IgEJOURNAL OF APPLIED TOXICOLOGY, Issue 4 2003R. J. Dearman Abstract No validated or widely recognized test methods are currently available for the prospective identi,cation of chemicals with the potential to cause respiratory allergy. The cellular and molecular mechanisms that result in the induction of chemical sensitization of the respiratory tract are unclear, although there is evidence for the selective development of T helper 2 (Th2)-type responses and, in some cases, the production of IgE antibody. We have therefore examined the utility of cytokine pro,ling using BALB/c mice, together with the measurement of induced increases in the total serum concentration of IgE in the Brown Norway (BN) rat, as markers for the prospective identi,cation of chemical respiratory allergens. Responses provoked by the reference respiratory allergen trimellitic anhydride (TMA) have been compared with those stimulated by the respiratory sensitizing diisocyanates toluene diisocyanate (TDI) and hexamethylene diisocyanate (HDI) and by the acid anhydride hexahydrophthalic anhydride (HHPA). Topical exposure of BN rats to TMA, TDI and HHPA each provoked marked immune activation (increases in lymph node cellularity and proliferation). However, only treatment with TMA stimulated vigorous increases in the total serum concentration of IgE. In contrast, exposure to HHPA, TDI or HDI failed to provoke signi,cant changes in serum IgE concentration or induced only transient and relatively weak increases in serum IgE levels. In parallel experiments using BALB/c strain mice, however, topical application of all four chemical respiratory allergens provoked a marked Th2-type cytokine secretion pro,le in draining lymph node cells. These data suggest that the measurement of induced changes in serum IgE is not suf,ciently sensitive for the robust identi,cation of chemical respiratory allergens. Furthermore, irrespective of the reasons for variations in TMA-induced IgE production among BN rats, doubts remain regarding the utility of these animals for the characterization of immune responses to chemical allergens. Cytokine pro,ling using the BALB/c strain mouse apparently provides a more robust method for the hazard assessment of chemical respiratory allergens. Copyright © 2003 John Wiley & Sons, Ltd. [source] | ||||||||||||||||||||||||||||||||||