Reed-Sternberg cells form by abscission failure in the presence of functional Aurora B kinase.

Large multinucleated Reed-Sternberg cells (RS) and large mononucleated Hodgkin cells (H) are traditionally considered to be the neoplastic population in classical Hodgkin lymphoma, (cHL) and postulated to promote the disease. However, the contribution of these larger cells to the progression of cHL remains debatable. We used established cHL cell lines and cHL cellular fractions composed of small mononucleated cells only or enriched in large RS/H cells to investigate RS/H cell origin and to characterize the cells which they derive from. We confirm that the small mononucleated cells give rise to RS/H cells, and we show that the latter proliferate significantly more slowly than the small cells. By using live-cell imaging, we demonstrate that binucleated RS cells are generated by failure of abscission when a few small cells attempt to divide. Finally, our results reveal that the small mononucleated cells are chromosomally unstable, but this is unlikely to be related to a malfunctioning chromosomal passenger protein complex. We propose that the small mononucleated cells, rather than the RS/H cells, are the main drivers of cHL.

The microtubule plus-end tracking protein CLASP2 is required for hematopoiesis and hematopoietic stem cell maintenance.

Mammalian CLASPs are microtubule plus-end tracking proteins whose essential function as regulators of microtubule behavior has been studied mainly in cultured cells. We show here that absence of murine CLASP2 in vivo results in thrombocytopenia, progressive anemia, and pancytopenia, due to defects in megakaryopoiesis, in erythropoiesis, and in the maintenance of hematopoietic stem cell activity. Furthermore, microtubule stability and organization are affected upon attachment of Clasp2 knockout hematopoietic stem-cell-enriched populations, and these cells do not home efficiently toward their bone marrow niche. Strikingly, CLASP2-deficient hematopoietic stem cells contain severely reduced mRNA levels of c-Mpl, which encodes the thrombopoietin receptor, an essential factor for megakaryopoiesis and hematopoietic stem cell maintenance. Our data suggest that thrombopoietin signaling is impaired in Clasp2 knockout mice. We propose that the CLASP2-mediated stabilization of microtubules is required for proper attachment, homing, and maintenance of hematopoietic stem cells and that this is necessary to sustain c-Mpl transcription.

U-2940, a human B-cell line derived from a diffuse large cell lymphoma sequential to Hodgkin lymphoma.

Several patterns of association between Hodgkin and non-Hodgkin lymphomas are recognized, some of which support a common cellular origin or shared transformation events for both malignancies. We describe the U-2940 cell line derived from a diffuse large B-cell lymphoma with some features consistent with mediastinal large B-cell lymphoma, clinically apparent 1 month after the initial course of chemotherapy for Hodgkin's disease, fulfilling the criteria for composite malignancies. U-2940 cells display a mature B phenotype with hypermutated IgH rearrangement typical of germinal/postgerminal center origin. The cell line is negative for Epstein-Barr virus and no evidence of t(14;18) was found. U-2940 cells display multiple chromosomal rearrangements similar to recurrent aberrations described in both Hodgkin and non-Hodgkin lymphomas, also partially shared by U-2932 derived from a B-cell lymphoma sequential to Hodgkin's disease. The original large B-cell lymphoma and the U-2940 cell line bear microsatellite instability, an abnormality associated with particular subtypes of non-Hodgkin lymphomas and found in tissues involved by Hodgkin lymphoma. Therefore, U-2940 cells bear several features known to occur in Hodgkin and in non-Hodgkin lymphomas, leading to the assumption that this cell line may constitute a useful tool to address elective pathways of lymphomagenesis and eventually the Hodgkin and non-Hodgkin lymphoma association.

Specific downregulation of bcl-2 and xIAP by RNAi enhances the effects of chemotherapeutic agents in MCF-7 human breast cancer cells.

Antiapoptotic genes such as bcl-2 or xIAP may be responsible for resistance to apoptosis induced by cytotoxic drugs. The aim of this study was to investigate if downregulation of bcl-2 or xIAP by RNA interference (RNAi) would sensitize MCF-7 cells to etoposide and doxorubicin. FITC-siRNAs uptake was verified by fluorescence microscopy and downregulation of Bcl-2 or XIAP was confirmed by Western Blotting. Both siRNAs reduced the number of viable cells and increased cellular apoptosis. Treatment with siRNAs followed by treatment with etoposide or doxorubicin further reduced the number of viable cells, when compared to either of the treatments alone. Therefore, downregulation of bcl-2 or xIAP by RNAi enhances the effects of etoposide and doxorubicin.

Survivin is required for stable checkpoint activation in taxol-treated HeLa cells.

Survivin is an essential chromosomal passenger protein whose function remains unclear. Here, we have used RNA interference to specifically repress Survivin in cultured HeLa cells. Immunoblot analysis showed that Survivin was no longer detectable in cultures 60 hours after transfection with Survivin-specific siRNA. Live cell analysis showed that many Survivin-depleted cells were delayed in mitosis, and immunofluorescence analysis of fixed specimens revealed that Survivin-depleted cells accumulated in prometaphase with misaligned chromosomes. The chromosomal passenger proteins, INCENP and Aurora-B, which can interact directly with Survivin, were absent from the centromeres of Survivin-depleted cells. These data contribute to the emerging picture that Survivin operates together with INCENP and Aurora-B to perform its mitotic duties. Some Survivin-depleted cells eventually exited mitosis without completing cytokinesis. This resulted in a gradual increase in the percentage of multinucleated cells in the culture. Time-lapse imaging of synchronized cultures revealed that control and Survivin-depleted cells arrested in mitosis in the presence of nocodazole; however, the latter failed to arrest in mitosis when treated with taxol. Immunofluorescence studies revealed that Survivin-depleted cells were unable to stably maintain BubR1 at the kinetochores in the presence of either taxol or nocodazole. Our data reveal that Survivin is not required for the spindle assembly checkpoint when it is activated by the loss of microtubules. However, Survivin is required for the maintenance of the checkpoint when it is activated by taxol, which is generally thought to cause a loss of spindle tension.

Poorly differentiated carcinomas of the thyroid gland: a review of the clinicopathologic features of a series of 28 cases of a heterogeneous, clinically aggressive group of thyroid tumors.

We evaluated the clinicopathologic, immunohistochemical, lectin histochemical, ultrastructural and morphometric characteristics of a series of 28 poorly differentiated carcinomas of the thyroid (PDCT). The 28 tumors were classified according to their predominant growth pattern as insular (n=12), trabecular (n=10) and solid subtypes (n=6). The overall mortality rate was 46%. No significant differences were found among the 3 subtypes. Data obtained by lectin histochemistry, electron microscopy, and morphometry point to a closer relationship of PDCT to follicular than to papillary carcinoma. Our results do not support the subdivision of PDCT into subtypes.

Hyalinizing trabecular adenoma: A misnomer for a peculiar tumor of the thyroid gland.

We describe the clinical, histological, immunohistochemical, and electron microscopical features of 9 tumors fulfilling the criteria of the so-called hyalinizing trabecular adenoma (HTA) of the thyroid. Six tumors had the characteristic histology of HTA throughout, whereas in the remaining 3 tumors the classic pattern was identified focally in otherwise typical or atypical follicular adenomas. In one case, there was a focus of tumoral tissue outside the capsule, and in another there was a regional lymph node metastasis. Seven tumors were immunoreactive for thyroglobulin and cytokeratins, 1 tumor was positive for thyroglobulin and negative for cytokeratins, and another was negative for thyroglobulin and positive for cytokeratins. Scattered cells immunoreactive for neurotensin and somatostatin were found in 2 cases. Every tumor stained for S100 protein and neuron-specific enolase, but none showed immunoreactivity for calcitonin, calcitonin gene-related peptide, or chromogranin. The irregularity of the nuclear contours, the prominence of the cytoplasmic bundles of intermediate filaments, and the accumulation of basal lamina material around the neoplastic cells without the interposition of a well-defined basal lamina were the most distinctive electron microscopical features. The cytogenetic study performed in one case revealed, apart from cells with a normal karyotype, two abnormal clones: one with a translocation of chromosomes 2-3 and another with the same translocation and trisomies of chromosomes 7 and 12. Our results show that most HTAs display follicular cell differentiation and very low clinical aggressiveness. They also show that some HTAs are able to coexpress follicular cell and neuroendocrine markers and may behave like malignant neoplasms. We conclude that hyalinizing trabecular tumor is a more appropriate generic term than HTA to designate this relatively heterogenous group of lesions of the thyroid gland.