Analysis of DNA distribution in Kaposi's sarcoma in patients with and without the acquired immune deficiency syndrome

While Kaposi's sarcoma in patients with the acquired immune deficiency syndrome (AIDS) may present as a multicentric disease with progressive organ involvement, the classic form of Kaposi's sarcoma is an indolent tumor seldom affecting extracutaneous areas and almost never responsible for the patient's demise. An attempt was made to correlate these clinical differences with the nuclear DNA content of tumor cells in histologic sections from 15 patients (9 with AIDS and 6 without AIDS). All tumors showed a similar DNA distribution pattern, with most cells appearing diploid, indicative of a low malignant potential. These findings indicate that Kaposi's sarcoma of both AIDS and non-AIDS patients is a tumor of intrinsically low malignancy and that lack of immune surveillance is most probably responsible for its aggressive biologic behavior in many AIDS patients.     

Spindle cells and their role in Kaposi's sarcoma

Spindle cells represent the main cell type of the advanced final nodular stage of Kaposi's sarcoma lesions. Despite some clinical and epidemiological differences, the four Kaposi's sarcoma forms (classic, endemic, post-transplant and epidemic) display very similar histopathological features, with the proliferation of spindle cells (considered as the Kaposi's sarcoma tumor cells) associated with inflammation and neo-angiogenesis. Electron-microscopy and immuno-histochemistry studies have led to the consensus that the spindle cells originated from the endothelial lineage. However, only recently, studies that used specific lymphatic immunological markers (such as podoplanin) and molecular features (gene expression microarrays) strongly linked Kaposi's sarcoma spindle cells to the endothelium lymphatic cell lineage. Both hybridization and immuno-histochemistry techniques have demonstrated that human herpesvirus 8 also known as Kaposi's sarcoma associated herpesvirus was present in spindle cells at all stages of the disease (patch, plaque, nodule). Interestingly, while the human herpesvirus 8 latent genes are expressed in nearly all tumor spindle cells, only a small fraction of them expresses markers of viral lytic replication. Recent findings showing that nodular Kaposi's sarcoma lesions display all patterns of human herpesvirus 8 clonality support the model according to which this tumor begins as a polyclonal disease with a subsequent evolution to a mono/oligoclonal process involving infected spindle cells. Spindle cells appear to be the central masterpiece in KS tumorigenesis, however the exact respective role of each human herpesvirus 8 gene, in the initiation and the disease progression is still under investigation and the question of whether or not this tumor is a reactive process or a true malignant proliferation of spindle cells remains yet unclear.     

Fine needle aspiration cytology of epithelioid angiosarcoma. Report of a case with nuclear grooves and indentations

BACKGROUND: Angiosarcoma, because of its rarity and histologic diversity, has been a persistent challenge to cytopathologists. Epithelioid angiosarcomas are often confused with carcinomas, melanomas and other epithelioid sarcomas, both cytologically and histologically. Here we report the cytopathologic features of a case of epithelioid angiosarcoma with prominent nuclear grooves. CASE: A 61-year-old male visited the hospital with a neck mass. On fine needle aspiration the hypercellular smear, with a lymphocytic bloody background, showed variable-sized, round to oval, pleomorphic cells with frequent nuclear grooves, indentations and plump cytoplasm. The cells were diffusely scattered or loosely aggregated, with occasional acinar and cell-in-cell configuration. The case was diagnosed as epithelioid angiosarcoma on excisional biopsy. CONCLUSION: Whenever fine needle aspiration cytology, especially in the head and neck area, suggests malignancy composed predominantly of epithelioid cells, epithelioid angiosarcoma should be considered. The differential diagnosis should include metastatic tumors, such as carcinoma, melanoma and primary sarcoma. Nuclear grooves and indentations are 1 of the important diagnostic features of epithelioid angiosarcoma.     

Differential expression of viral Bcl-2 encoded by Kaposi's sarcoma-associated herpesvirus and human Bcl-2 in primary effusion lymphoma cells and Kaposi's sarcoma lesions

Expression of human herpesvirus 8 viral Bcl-2 protein was demonstrated in spindle cells of late-stage Kaposi's sarcoma lesions but not in primary effusion lymphoma cell lines. In contrast, strong expression of human Bcl-2 was found in stimulated primary effusion lymphoma cells, whereas in Kaposi's sarcoma lesions preferential mononuclear cells, and to a lesser extent spindle cells, stained positive.     

Cytomorphologic features of angiosarcoma on fine needle aspiration biopsy.

OBJECTIVE: To present the cytomorphologic features of angiosarcomas identified on fine needle aspiration biopsy, review the literature, and discuss the differential diagnosis and pitfalls involved in such cases. STUDY DESIGN: Fine needle aspirate smears from 11 cases (1 hepatic, 3 breast and 7 subcutaneous/soft tissue lesions of angiosarcomas from eight patients were reviewed. All cases had histologic confirmation of angiosarcoma. RESULTS: All aspirates were hypocellular, with predominantly single cells in a background of moderate to abundant amounts of blood. Nine cases had scattered inflammatory cells, primarily neutrophils, in the background. Six of the cases had rare small clusters of cells. The cells were oval, round or spindled, with eccentric, round to spindle-shaped nuclei and moderate to abundant amounts of pale blue-gray, vacuolated cytoplasm. The cells ranged from two to nine times the size of the background red blood cells. In four cases, malignant cells demonstrated intracytoplasmic hemosiderin deposits. Small nucleoli were identified in five cases, large nucleoli in one case and hyperchromasia in seven cases. Mitotic figures, erythrophagocytosis, acinarlike or vascular structures, and necrosis were not identified in any of the studied cases. In four cases, a definitive diagnosis of angiosarcoma was rendered on the fine needle aspiration specimen. In three other cases, the differential diagnosis remained between angiosarcoma and radiation change. CONCLUSION: The presence of scarce single pleomorphic cells in a bloody background should raise the diagnostic possibility of angiosarcoma. A definitive diagnosis of angiosarcoma is often difficult to render due to the paucity of diagnostic cells unless intracytoplasmic hemosiderin deposits can be identified. Multiple aspirations are often needed in order to obtain diagnostic material. In the setting of radiotherapy, it may be impossible to distinguish angiosarcoma from radiation change, and biopsy should be recommended.     

Superficial urothelial (umbrella) cells. A potential cause of abnormal DNA ploidy results in urine specimens

OBJECTIVE: To determine the DNA ploidy distribution in urothelial superficial (umbrella) cells and to assess the value of the image analysis operator's experience. STUDY DESIGN: DNA ploidy was assessed in 12 cytologically negative bladder washes stained with Feulgen stain. All 12 cases were evaluated independently by three operators with different levels of cytopathology experience and different goals. Operator 1 (experienced) selected only nuclei of urothelial cells, avoiding nuclei of superficial cells; operator 2 (experienced) selected only nuclei of superficial cells; operator 3 (inexperienced) selected the largest and most-atypical-looking nuclei. Each operator measured a total of 100 nuclei per case. RESULTS: Operator 1 found all cases to be diploid (97% of nuclei on average). Operators 2 and 3 showed a wide range of results. Almost half the nuclei (47%) analyzed by operator 2 were in the diploid region, a third (35%) were in the tetraploid region, and the remaining (18%) ones had a DNA index (DI) in the range of 1.2-1.8 or > 2.5. Operator 3 obtained the most abnormal results. Only 9% of the nuclei were diploid, while 37% were in the tetraploid region, 18% were in the hyperploid region, and 35% had a DI in the range of 1.2-1.8. Differences among results obtained by each operator were statistically significant. CONCLUSION: The nuclei of superficial (umbrella) cells often have abnormal DNA content, which may cause abnormal DNA ploidy results in cytomorphologically normal bladder washes. Consequently, the nuclei of superficial cells should be avoided in the evaluation of urine samples. DNA analysis of urine specimens requires selection of nuclei only of deep urothelial cells by an experienced operator.     

Angiosarcoma in FNA smears: diagnostic accuracy,morphology, immunocytochemistry and differential diagnoses

?. Pohar‐Marin?ek and J. Lamovec Angiosarcoma in FNA smears: diagnostic accuracy, morphology, immunocytochemistry and differential diagnoses Objective: The aim of our study was to analyse the diagnostic accuracy in recognizing angiosarcoma from fine needle aspiration (FNA) samples and to determine morphological features of angiosarcoma in cytology. Methods: FNA samples from 18 histologically confirmed angiosarcomas obtained between 1985 and 2009 were included in the study. Original cytological diagnoses were retrieved, smears reviewed and morphological features analysed: cellularity, smear pattern, cell morphology, contents of background. Outcome of immunocytochemistry was noted and additional reactions performed if material was available. Results: There were 13 primary angiosarcomas and five recurrent tumours; nine tumours were epithelioid. Twelve tumours were cytologically diagnosed as malignant, three as suspicious and three were judged unsatisfactory. Only two primary tumours were diagnosed as vascular. According to morphology, tumours were divided into those with predominantly epithelioid cells and those with predominantly spindle cells. Within these two groups were variations due to grade of tumour. Cytomorphology did not correlate well with histology in mixed and spindle cell types of angiosarcomas. Immunocytochemistry was applied in seven cases, specific vascular marker CD31 only twice at the time of diagnosis and three times retrospectively. Conclusions: Angiosarcomas are difficult to recognize on FNA smears when they lack the typical dual, spindle and epithelioid cell population and when they occur in internal organs where carcinomas are more common. Very few reliable data are available concerning specificity of CD31 on cytological material.     

NKG2C+ NK cells are enriched in AIDS patients with advanced-stage Kaposi's sarcoma

Kaposi's sarcoma (KS) is an AIDS-defining condition in individuals with human immunodeficiency virus type 1 infection. We investigated the phenotype and function of the NKG2C+ NK cell population in individuals with AIDS and Kaposi's sarcoma. The staging of AIDS KS patients according to the AIDS Clinical Trial Group criteria revealed that patients with the S1 disease stage have a significantly higher proportion of NKG2C+ cells than those with the S0 disease stage. NKG2C+ cells from S1-stage patients are highly enriched for the expression of KIR3DL1, are depleted of NKp46, and respond poorly to major histocompatibility complex class I-positive target cells. These data demonstrate a link between NK cell phenotype and function and disease prognosis in AIDS.     

Activation of NF-kappaB by the latent vFLIP gene of Kaposi's sarcoma-associated herpesvirus is required for the spindle shape of virus-infected endothelial cells and contributes to their proinflammatory phenotype

Kaposi's sarcoma (KS) is an inflammatory angioproliferative lesion induced by the infection of endothelial cells with the KS-associated herpesvirus (KSHV). Infected endothelial cells assume an elongated (spindle) shape that is one of the histologic signatures of KS. In vitro, latent viral infection of primary endothelial cells (but no other cell type) strikingly recapitulates these morphological findings. Here we report that the spindling phenotype involves major rearrangement of the actin cytoskeleton and can be attributed to the expression of a single viral protein, vFLIP, a known activator of NF-kappaB. Consistent with this, the inhibition of NF-kappaB activation blocks vFLIP-induced spindling in cultured endothelial cells. vFLIP expression in spindle cells also induces the production of a variety of proinflammatory cytokines and cell surface adhesion proteins that likely contribute to the inflammatory component of KS lesions.     

Cloning and characterization of murine EDF-1

Murine endothelial differentiation-related factor (mEDF-1) encodes a basic intracellular protein of 148 amino acids which is highly homologous to the human and rat polypeptides. mEDF-1 is expressed in most murine tissues tested and is evolutionary conserved. mEDF-1 expression is modulated in mouse development, since its expression is high early in development and decreases thereafter. Because EDF-1 has been isolated as a gene differentially expressed by exposure of endothelial cells to the Tat protein of HIV, we evaluated mEDF-1 expression in different cell lines derived from tumors which spontaneously develop in Tat transgenic mice. Cells isolated from adenocarcinomas and leiomyosarcomas express very high amounts of EDF-1, independently from their capability to secrete Tat. Tat transgenic mice also develop skin lesions which closely resemble human Kaposi's sarcoma. Since Kaposi spindle cells, which are the proliferative component of the sarcoma, differentiate from an endothelial precursor, it is noteworthy that spindle cells derived from Kaposi-like lesions of the Tat transgenic mice downregulate EDF-1 when compared to microvascular endothelial cells isolated from the same tissue.     

Photoradiation therapy of cutaneous angiosarcomas in mice

Cutaneous angiosarcoma is a relatively rare but devastating malignant vascular tumor. It has a high incidence of recurrence following conventional therapeutic modalities applied either singly or in combination. The increased vascularity of cutaneous angiosarcomas, facilitating selective uptake and retention of a photosensitizing agent, such as hematoporphyrin derivative (HPD), suggests that these tumors would respond well to photoradiation therapy. To study the feasibility of this treatment modality, transplantable hemangiosarcomas were implanted in B6C3F1 female mice. Within 2.5 to 3.5 hours after intraperitoneal administration of HPD, fluorescence was recorded in the tumor as compared with surrounding normal skin. When these photosensitized tumors were exposed to 70 J/cm2 of laser energy from an argon-pumped dye laser at 630 nm, the tumors showed marked necrosis within 24 hours. In another series, the tumors were initially photosensitized with HPD for 3 hours and then treated with laser energy ranging from 0 to 96 J/cm2. A dual labeling procedure demonstrated a dose-related decrease in DNA synthesis rate in tumors that were exposed to 0 to 30 J/cm2 at 24 hours after treatment. Furthermore, tumor tissue exposed to laser energy in excess of 30 J/cm2 showed no significant cellular DNA synthesis. These data, supported by histologic evidence of tissue destruction, suggest that photoradiation therapy has a great potential as a therapeutic modality for cutaneous angiosarcomas.     

Mechanism of paclitaxel activity in Kaposi's sarcoma

Kaposi's sarcoma (KS) is an angioproliferative disease characterized by proliferation of spindle-shaped cells predominantly of endothelial cell origin, neoangiogenesis, inflammatory cell infiltration, and edema. At least in early stage, KS behaves as a reactive lesion sustained by the action of inflammatory cytokines and growth factors, has a polyclonal nature, and can regress. However, in time it can become monoclonal, especially in the nodular stage, evolving into a true sarcoma, likely in association with the increased expression of antiapoptotic oncogenes. We have recently demonstrated by immunohistochemical analysis that Bcl-2, a proto-oncogene known to prolong cellular viability and to antagonize apoptosis, is highly expressed in spindle cells and vessels of both AIDS-KS and classical KS lesions and that its expression increases with lesion stage. Paclitaxel, a microtubule-stabilizing drug known to inhibit Bcl-2 antiapoptotic activity and to be highly effective in the treatment of certain neoplasms, has recently been found to be active also in patients with advanced HIV-associated KS. In this report we investigated the mechanism(s) of paclitaxel activity in KS. By using a model of experimental KS induced by the inoculation of KS-derived spindle cells in nude mice and primary cultures of KS spindle cells, we found that paclitaxel promotes regression of KS lesions in vivo and that it blocks the growth, migration, and invasion of KS cells in vitro. Furthermore, paclitaxel treatment promoted apoptosis and down-regulated Bcl-2 protein expression in KS cells in vitro and in KS-like lesions in mice. Our results suggest that paclitaxel interferes with KS by down-regulating Bcl-2 antiapoptotic effect.     

PML down‐regulation in soft tissue sarcomas

To date, little is known concerning the promyelocytic leukemia gene (PML) status in tumors of different origin, and its expression has never been evaluated in soft tissue sarcoma. The aim of the present study is focused on the identification of differences in terms of PML protein expression between different types of soft tissue sarcoma and the corresponding normal surrounding tissue. PML protein expression has been assessed by immunohistochemistry in six different histologic types of soft tissue sarcoma (synovial sarcoma, myofibroblastic sarcoma, angiosarcoma, liposarcoma, pleomorphic sarcoma, and leiomyosarcoma) and in the corresponding normal surrounding tissue. PML resulted significantly down‐regulated in synovial sarcoma and in myofibroblastic sarcoma specimens. Also in angiosarcoma samples a significative difference in PML expression in comparison with normal specimens has been detected. Interestingly PML protein detection showed a different pattern of expression in the three liposarcoma histology types compared with corresponding nontumoral tissues. In particular PML protein resulted significantly down‐regulated in myxoid liposarcoma and in dedifferentiated liposarcoma. On the contrary no statistically significant difference was observed in pleomorphic liposarcoma compared to normal tissue specimens. Further investigations are needed to confirm these data and to assess the possible value of PML expression as a prognostic factor in these extremely aggressive diseases. J. Cell. Physiol. 224: 644–648, 2010. © 2010 Wiley‐Liss, Inc.     

人疱疹病毒8型KS330基因片段的检出与Kaposi肉瘤的关系

HHV-8 sequences were recently identified in 100% of the amplifiable samples from AIDS patients with Kaposi's sarcoma(KS)and in 15% of the non-KS tissue samples from AIDS patients, so there is a strong correlation of Kaposi's sarcoma with HHV-8. Serum and DNA samples from a clinically diagnosed Kaposi's sarcoma Chinese patient were tested. HHV-8 antibody was tested positive by IFA and HIV-I antibody was negative by Western blot. The KS330 PCR product was found both in peripheral blood mononuclear cells and in KS tumor cells from this Chinese patient. This supports the hypothesis that Kaposi's sarcoma results from infection of HHV-8.     

Quantitative DNA analysis of low grade cervical intraepithelial neoplasia and human papillomavirus infection by static and flow cytometry.

Quantitative deoxyribonucleic acid (DNA) analysis of cervical biopsy specimens from 26 women with cytological, colposcopic, and histological evidence of mild cervical atypia consistent with cervical intraepithelial neoplasia grade I, reactive atypia, or human papillomavirus infection alone or in combination was performed in a comparative evaluation of Feulgen microspectrophotometry, the fast interval processor image analysis system, and flow cytometry. The fast interval processor image analysis system showed a distinct advantage over the other methods, being faster and allowing the operator to see the cells that were selected for measurement. The three methods of measurement together showed that the DNA content of at least 2% of the cells measured exceeded 5C (C being the haploid amount of DNA in a normal cell and 2C representing the diploid complement of a normal cell) in all cases of cervical intraepithelial neoplasia grade I and reactive atypia and in 87% of those reported as showing human papillomavirus infection alone. In contrast, the DNA content of cervical biopsy specimens from the transformation zone of 11 normal controls did not exceed 4C. This study shows the value of using a DNA threshold--that is, the "5C exceeding rate"--to distinguish between normal and neoplastic appearances of the cervix. These results support the view that cervical infection by human papillomavirus is a true precursor of neoplasia.