In-vitro model for the ultrastructural study of the formation of thrombi in human platelets.

Platelets are cell fragments with dynamic properties involved in clot formation after tissue damage. Platelet activation causes a change in shape, secretion of intracellular granules and aggregation with each other through the cytoskeleton components and biochemical changes. Platelet adhesion, considered as the major event in haemostasis, has been studied in several in-vitro and in-vivo models to evaluate the feasible thrombogenicity of some materials, the dynamics of specific receptors, as well as the effect of different buffers and inhibitors in this process. In spite of the numerous reports about platelet activation, to date there is no information available about the fine structure of the platelet-platelet and platelet-substrate interactions. In the present report we describe an in-vitro system that allows the visualization of these interactions: platelets are adhered to an inert substrate, and interactions with suspended platelets as a process to initiate the formation of thrombi was followed by ultramicrotomy and transmission electron microscopy.     

18F-FDG PET/CT delayed images after diuretic for restaging invasive bladder cancer.

PET with (18)F-FDG has been considered of limited value for detection of bladder cancer because of the urinary excretion of the tracer. The purpose of this study was to investigate the role of PET/CT in the detection and restaging of bladder cancer using furosemide and oral hydration to remove the excreted (18)F-FDG from the bladder. METHODS: Seventeen patients with bladder cancer (11 without cystectomy, 6 with total cystectomy and urinary diversion) underwent (18)F-FDG PET/CT from head to the upper thighs 60 min after the intravenous injection of 370 MBq of (18)F-FDG. Additional pelvic images were acquired 1 h after the intravenous injection of furosemide and oral hydration. PET/CT findings were confirmed by MRI, cystoscopy, or biopsy. RESULTS: PET/CT was able to detect bladder lesions in 6 of 11 patients who had not undergone cystectomy. These images changed the PET/CT final reading in 7 patients: Recurrent bladder lesions were detected in 6 patients, pelvic lymph node metastases in 2 patients, and prostate metastasis in 1. This technique overcame the difficulties posed by the urinary excretion of (18)F-FDG. Hypermetabolic lesions could be easily detected by PET and precisely localized in the bladder wall, pelvic lymph nodes, or prostate by CT. Seven of 17 patients (41%) were upstaged only after delayed pelvic images. CONCLUSION: Detection of locally recurrent or residual bladder tumors can be dramatically improved using (18)F-FDG PET/CT with delayed images after a diuretic and oral hydration.     

Human papillomavirus detection: verification with cervical cytology.

OBJECTIVE: Thirteen specific types of human papillomavirus (HPV), classified as high-risk for the development of cervical cancer, have been reported in 99.7% of all cervical cancers. For this reason, and because of the reported lack of sensitivity of the Papanicolaou (Pap) smear for detecting HPV, some experts believe that the use of papillomavirus DNA testing may replace cytology for routine gynecological screening. Our goal was to validate a commercial assay, the Digene Hybrid Capture-2 for the detection of human papillomavirus by comparing the results to cytological detection of cervical abnormalities. DESIGN: Cytology results of concurrent liquid-based Papanicolaou smears were compared to the Hybrid Capture-2 results. Correlation was assessed and discordant cytology results were reviewed. SETTING: Louisiana State University Health Sciences Center at Shreveport, Department of Pathology, HPV Diagnostic Laboratory. PATIENTS: All liquid cytology specimens submitted for HPV testing between November 1, 2000 and April 1, 2001. RESULTS: Of the 291 cases tested by Hybrid Capture-2, 12% and 28% were positive with the low-risk and high-risk probes, respectively, and 265 had concurrent cytology results. Fourteen specimens testing positive only with the low-risk probe were not included in this comparison. Thus, the results for 251 of the 291 (86%) specimens tested for human papillomavirus DNA were compared to the original cytology report. Overall concordance between Hybrid Capture-2 and the original smear cytology result was 78%. Slide review reduced the number of discordant specimens from 22% to 12%. CONCLUSION: Based upon these data, we find the HPV assay to be useful as a routine screen for Human papillomavirus.     

Electron microscopy renders the diagnostic capabilities of cytopathology more precise: an approach to everyday practice

Cytology is a powerful diagnostic tool but to make definitive diagnoses, the use of ancillary techniques is imperative. By combining immunohistochemistry (IHC) and electron microscopy (EM), cytologic diagnoses can be as precise as those of surgical pathology. In the authors' daily practice of cytopathology they use all ancillary techniques available to them: histochemistry, IHC, EM, flow cytometry, and molecular pathology. IHC is frequently used as an ancillary technique in their daily practice but EM is many times their technique of choice. By the use of EM the authors can make specific final diagnoses, make the diagnosis more definitive, narrow the differential diagnosis, or determine the origin of a neoplasm with unknown primary site. Specimens obtained by fine-needle aspiration as well as all body fluids are suitable for EM. The limiting factor is to obtain the appropriate material with the diagnostic cells for ultrastructural examination. The common diagnostic dilemmas in the everyday practice of cytology are the following: mesothelioma vs. adenocarcinoma, neuroendocrine differentiation or not, the distinction of melanoma from adenocarcinoma and sarcoma, hepatocellular carcinoma vs. adenocarcinoma, and the origin of adenocarcinomas of unknown primary. The authors discuss how they approach these diagnostic problems in their everyday practice and how they incorporate EM in solving them.     

Comparison of the neuropsychological assessment in a girl with bilateral cerebrovascular disease (moyamoya) before and after surgical intervention

Moyamoya is a chronic progressive cerebrovascular disease with characteristic angiographic findings and a clinical picture with episodes of transient ischemic attacks, headache, seizures, hemiparesis, which may resolve after surgical treatment. We describe the case of a girl with the typical findings of the disease, comparing them before and after surgery with the use of neuropsychological tests, neurological examination and laboratory tests.     

Tc-99m MDP uptake in uterine leiomyoma

A 47-year-old woman with adenocarcinoma of the right breast had bone scintigraphy with Tc-99m MDP. Bone imaging did not show any metastases. However, a large area of increased tracer uptake was seen extending from the abdomen to the pelvis. Abdominal ultrasound revealed a large solid and heterogeneous mass, measuring 18 x 11 x 14.3 cm, that originated in an empty uterus. A biopsy of the surgical specimen showed a leiomyoma of the uterus.     

Organ slice viability extended for pathway characterization: an in vitro model to investigate fibrosis.

Liver slice viability is extended to 96 h for rat, expanding the use of this in vitro model for studying mechanisms of injury and repair, including pathways of fibrosis. The contributing factors to increased organ slice survival consist of the use of a preservation solution for liver perfusion and slice preparation, obtaining rats that are within the weight range of 250-325 g, placing a cellulose filter atop the titanium mesh roller-insert to support the slice, and maintaining the slices in an optimized culture medium which is replaced daily. The liver slices remain metabolically active, synthesizing adenosine triphosphate (ATP), glutathione, and glycogen, and exhibit preserved organelle integrity and slice morphology. Slice preparation results in 2-cut surfaces which likely triggers a repair and regenerative response. The fibrogenic pathways are evident by the activation of stellate cells, the proliferation of myofibroblast-like cells, and an increased collagen deposition by 48 h. Markers indicative of activated stellate cells, alpha-smooth muscle actin, collagen 1a1, desmin, and HSP47 are substantiated by real time-PCR. Increased staining of alpha-smooth muscle actin initially around the vessels and by 72-96 h in the tissue is accompanied by increased collagen staining. Microarray gene expression revealed extracellular matrix changes with the up-regulation of cytoskeleton, filaments, collagens, and actin genes; and the down-regulation of genes linked with lipid metabolism. The improvements in extending liver slice survival, in conjunction with its three-dimensional multi-cellular complexity, increases the application of this in vitro model for investigating pathways of injury and repair, and fibrosis.     

Role of Intracystic Cytokines and Nitric Oxide in Ovarian Neoplasms

The development of new biomarkers for the diagnosis and prognosis of ovarian cancer may provide an opportunity for new therapies. In this study, we aimed to compare cytokines (interleukin [IL]‐2, IL‐5, IL‐6, IL‐8, IL‐10 and tumour necrosis factor [TNF]‐α) and nitric oxide (NO) metabolite levels in non‐neoplastic tumours, benign primary ovarian tumours and malignant primary ovarian neoplasms. The secondary aim was to relate cytokine and intracystic NO metabolite levels to clinical, laboratory and pathologic characteristics for patients with primary ovarian malignancies. We evaluated 110 patients with adnexal masses. Cytokine concentrations were quantified by enzyme‐linked immunosorbent assay and nitrate concentrations by enzymatic reduction of nitrite by nitrate reductase. Patients with malignant neoplasms had higher IL‐6, IL‐8 and NO levels compared to patients with benign neoplasms. Histologic grade 1 tumours were associated with elevated IL‐2 levels, whereas anaemia was associated with elevated IL‐6 levels. On average, those patients with elevated IL‐8 levels also had a neutrophil/lymphocyte ratio (NLR) greater than 2.6 and less than 36 months of disease‐free survival (DFS). Patients with normal CA 19‐9 levels had elevated IL‐10 levels. TNF‐α was elevated in patients with two carcinogenesis and those with a platelet/lymphocyte ratio (PLR) less than 300. NO levels were higher in patients with an NLR less than 2.6 and CA 19‐9 greater than 35 U/ml. Elevated intracystic cytokine levels, especially IL‐6 and IL‐8, are associated with worse prognosis in ovarian cancer.     

Electron Microscopy Renders the Diagnostic Capabilities of Cytopathology More Precise: An Approach to Everyday Practice

Cytology is a powerful diagnostic tool but to make definitive diagnoses, the use of ancillary techniques is imperative. By combining immunohistochemistry (IHC) and electron microscopy (EM), cytologic diagnoses can be as precise as those of surgical pathology. In the authors' daily practice of cytopathology they use all ancillary techniques available to them: histochemistry, IHC, EM, flow cytometry, and molecular pathology. IHC is frequently used as an ancillary technique in their daily practice but EM is many times their technique of choice. By the use of EM the authors can make specific final diagnoses, make the diagnosis more definitive, narrow the differential diagnosis, or determine the origin of a neoplasm with unknown primary site. Specimens obtained by fine-needle aspiration as well as all body fluids are suitable for EM. The limiting factor is to obtain the appropriate material with the diagnostic cells for ultrastructural examination. The common diagnostic dilemmas in the everyday practice of cytology are the following: mesothelioma vs. adenocarcinoma, neuroendocrine differentiation or not, the distinction of melanoma from adenocarcinoma and sarcoma, hepatocellular carcinoma vs. adenocarcinoma, and the origin of adenocarcinomas of unknown primary. The authors discuss how they approach these diagnostic problems in their everyday practice and how they incorporate EM in solving them.     

α-Fetoprotein,tissue polypeptide antigen and ferritin in diagnosing primary hepatocellular carcinoma in patients with liver cirrhosis

Summary This study was undertaken in order to compare the usefulness of three indices of tumour proliferation in detecting primary hepatocellular carcinoma (HCC) and in differentiating this neoplasm from liver cirrhosis. In 10 patients with HCC and in 63 with liver cirrhosis serum -fetoprotein (AFP), tissue polypeptide antigen (TPA) and ferritin were assayed. Increased levels of AFP but not of TPA and ferritin were observed in HCC as compared to liver cirrhosis. The receiver-operating characteristic curves demonstrated that AFP is more discriminating between HCC and liver cirrhosis than the other two markers. Correlations between liver function tests and serum markers were observed in liver cirrhosis but not in HCC. We can conclude that AFP is more useful than TPA and ferritin in detecting HCC in patients with liver cirrhosis, owing to the high frequency of false positive results of the latter two indices in liver cirrhosis. Liver dysfunction is probably involved in increasing all these markers of malignancy, thus reducing the specificity of these tests.Abbreviations AFP -tetoprotein - TPA tissue polypeptide antigen - HCC primary hepatocellular carcinoma Partially supported by a grant of the Italian National Research Council, Special Project Oncology, contract 87.01.541.04. Under the auspices of the R. Farini association for gastroenterological research