Burkitt-like lymphoma with 11q aberration: A germinal center derived lymphoma genetically unrelated to Burkitt lymphoma

Burkitt-like lymphoma with 11q aberration is characterized by pathological features and gene expression profile resembling Burkitt lymphoma but lack MYC rearrangement and carries an 11q-arm aberration with proximal gains and telomeric losses. Whether these lymphomas are a distinct category or a particular variant of other recognized entities is controversial. To improve the understanding of Burkitt-like lymphoma with 11q aberration we have performed an analysis of copy number alterations and targeted sequencing of a large panel of B-cell lymphoma related genes in 11 cases. Most patients had localized nodal disease and a favourable outcome after therapy. Histologically, they were high grade B-cell lymphoma, not otherwise specified (8 cases), diffuse large B-cell lymphoma (2 cases) and only one was considered as atypical Burkitt lymphoma. All cases had a germinal center B-cell signature and phenotype with frequent LMO2 expression. Burkitt-like lymphoma with 11q aberration had frequent gains of 12q12-q21.1 and losses of 6q12.1-q21, and lacked common Burkitt lymphoma or diffuse large B-cell lymphoma alterations. Potential driver mutations were found in 27 genes, particularly involving BTG2, DDX3X, ETS1, EP300, and GNA13. However, ID3, TCF3, or CCND3 mutations were absent in all cases. These results suggest that Burkitt-like lymphoma with 11q aberration is a germinal center derived lymphoma closer to high grade B-cell lymphoma or diffuse large B-cell lymphoma rather than Burkitt lymphoma.Copyright © 2019, Ferrata Storti Foundation

N-Terminal Prolactin-Derived Fragments, Vasoinhibins, Are Proapoptoptic and Antiproliferative in the Anterior Pituitary

The anterior pituitary is under a constant cell turnover modulated by gonadal steroids. In the rat, an increase in the rate of apoptosis occurs at proestrus whereas a peak of proliferation takes place at estrus. At proestrus, concomitant with the maximum rate of apoptosis, a peak in circulating levels of prolactin is observed. Prolactin can be cleaved to different N-terminal fragments, vasoinhibins, which are proapoptotic and antiproliferative factors for endothelial cells. It was reported that a 16 kDa vasoinhibin is produced in the rat anterior pituitary by cathepsin D. In the present study we investigated the anterior pituitary production of N-terminal prolactin-derived fragments along the estrous cycle and the involvement of estrogens in this process. In addition, we studied the effects of a recombinant vasoinhibin, 16 kDa prolactin, on anterior pituitary apoptosis and proliferation. We observed by Western Blot that N-terminal prolactin-derived fragments production in the anterior pituitary was higher at proestrus with respect to diestrus and that the content and release of these prolactin forms from anterior pituitary cells in culture were increased by estradiol. A recombinant preparation of 16 kDa prolactin induced apoptosis (determined by TUNEL assay and flow cytometry) of cultured anterior pituitary cells and lactotropes from ovariectomized rats only in the presence of estradiol, as previously reported for other proapoptotic factors in the anterior pituitary. In addition, 16 kDa prolactin decreased forskolin-induced proliferation (evaluated by BrdU incorporation) of rat total anterior pituitary cells and lactotropes in culture and decreased the proportion of cells in S-phase of the cell cycle (determined by flow cytometry). In conclusion, our study indicates that the anterior pituitary production of 16 kDa prolactin is variable along the estrous cycle and increased by estrogens. The antiproliferative and estradiol-dependent proapoptotic actions of this vasoinhibin may be involved in the control of anterior pituitary cell renewal

Diverse mutations and structural variations contribute to Notch signaling deregulation in paediatric T-cell lymphoblastic lymphoma

Background T-cell lymphoblastic lymphoma (T-LBL) is an aggressive neoplasm closely related to T-cell acute lymphoblastic leukaemia (T-ALL). Despite their similarities, and contrary to T-ALL, studies on paediatric T-LBL are scarce and, therefore, its molecular landscape has not yet been fully elucidated. Thus, the aims of this study were to characterize the genetic and molecular heterogeneity of paediatric T-LBL and to evaluate novel molecular markers differentiating this entity from T-ALL. Procedure Thirty-three paediatric T-LBL patients were analyzed using an integrated approach, including targeted next-generation sequencing, RNA-sequencing transcriptome analysis and copy-number arrays. Results Copy number and mutational analyses allowed the detection of recurrent homozygous deletions of 9p/CDKN2A (78%), trisomy 20 (19%) and gains of 17q24-q25 (16%), as well as frequent mutations of NOTCH1 (62%), followed by the BCL11B (23%), WT1 (19%) and FBXW7, PHF6 and RPL10 genes (15%, respectively). This genetic profile did not differ from that described in T-ALL in terms of mutation incidence and global genomic complexity level, but unveiled virtually exclusive 17q25 gains and trisomy 20 in T-LBL. Additionally, we identified novel gene fusions in paediatric T-LBL, including NOTCH1-IKZF2, RNGTT-SNAP91 and DDX3X-MLLT10, the last being the only one previously described in T-ALL. Moreover, clinical correlations highlighted the presence of Notch pathway alterations as a factor related to favourable outcome. Conclusions In summary, the genomic landscape of paediatric T-LBL is similar to that observed in T-ALL, and Notch signaling pathway deregulation remains the cornerstone in its pathogenesis, including not only mutations but fusion genes targeting NOTCH1.We thank the centres of the Sociedad Espanola de Hematologia y Oncologia Pediatricas that submitted cases for consultation, to Noelia Garcia, Silvia Martin and Helena Suarez for their excellent technical assistance and to Nerea Dominguez for updating clinical data. We are indebted to the IDIBAPS Genomics Core Facility and to the HCB-IDIBAPS, the HospitaI Infantil Sant Joan de Deu and the Hospital Universitari Vall d'Hebron Tumour Biobanks, all integrated in the National Network Biobanks of ISCIII for the sample and data procurement. This work was supported by Asociacion Espanola Contra el Cancer (AECC CICPFI6025SALA and 'Ayudas Clinico Formacion AECC 2020' to Jaime Verdu-Amoros), Asociacion de aitas y amas para la humanizacion, socializacion e investigacion del Cancer Infantil y la divulgacion de la donacion de medula osea-La Cuadri del Hospi, Fondo de Investigaciones Sanitarias Instituto de Salud Carlos III (Miguel Servet Program I and II CP13/00159 and MSII18/00015; Itziar Salaverria), Generalitat de Catalunya Suport Grups de Recerca (2017-SGR-1107; Itziar Salaverria), and the European Regional Development Fund 'Una manera de fer Europa'. Joan Enric Ramis-Zaldivar was supported by a fellowship AGAUR FI-DGR 2017 (2017 FI_B01004) from Generalitat de Catalunya. Noelia Garcia has been continuously supported by Accio instrumental d'incorporacio de cientifics i tecnlegs PERIS 2016 (SLT002/16/00336) and PERIS 2020 (SL017/20/000204) from Generalitat de Catalunya. Julia Salmeron-Villalobos was supported by a fellowship from La Caixa (CLLEvolution-HR17-00221). This work was developed partially at the Centro Esther Koplowitz, Barcelona, Spain

Tumor Necrosis Factor-alpha induces apoptosis of lactotropes from female rats

TNF-α is involved in the regulation of normal tissue homeostasis affecting cell proliferation, differentiation, and death. We previously reported that TNF-α reduces anterior pituitary cell proliferation and PRL release in an estrogen-dependent manner. In the present project we studied the induction of apoptosis by TNF-α in anterior pituitary cells from female rats. TNF-α (50 ng/ml) decreased the viability of anterior pituitary cells. Incubation with TNF-α for 24 h increased the percentage of terminal deoxynucleotidyltransferase-mediated deoxyuridine triphosphate nick end labeling-positive cells. TNF-α increased the percentage of somatotropes and lactotropes with apoptotic nuclear morphology without affecting the proportion of apoptotic corticotropes or gonadotropes. TNF-α increased the percentage of apoptotic lactotropes in cultured cells from rats killed in proestrus and estrus, but not in diestrus. This effect was significantly higher in cells from rats in proestrus than in estrus. In anterior pituitary cells from ovariectomized rats, TNF-α significantly increased the percentage of apoptotic lactotropes only when the cells were incubated in the presence of 17β-estradiol. These results indicate that TNF-α induces apoptosis in somatotropes and lactotropes from female rats. The apoptotic effect of TNF-α on lactotropes is dependent on estrogens and could be involved in the regulation of anterior pituitary cell renewal during the estrous cycle.Fil: Candolfi, Marianela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Zaldivar, Verónica. Universidad de Buenos Aires. Facultad de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: de Laurentiis, Andrea. Universidad de Buenos Aires. Facultad de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Jaita, Gabriela Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Pisera, Daniel Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Seilicovich, Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentin

Nitric oxide mediates the inhibitory effect of tumor necrosis factor-α on prolactin release

Tumor necrosis factor-α (TNF-α) is a pleiotropic cytokine that markedly affects neuroendocrine functions. This cytokine is expressed in the anterior pituitary where its receptors are also present. Nitric oxide (NO) is synthesized in gonadotropes and folliculo-stellate cells of the anterior pituitary. Since NO directly inhibits prolactin secretion, we investigated the involvement of NO in the inhibitory effect of TNF-α on prolactin release from anterior pituitary cells of female rats. The presence of L-NAME (1 mM), an inhibitor of NO synthase (NOS), in the incubation medium significantly blunted the inhibition of prolactin release produced by TNF-α (50 ng/ml). TNF-α increased nitrite release to the incubation medium. The activity of NOS as measured by [14C]citrulline production was significantly enhanced when anterior pituitary cells were incubated with TNF-α for 8 h or more. Also, TNF-α induced iNOS gene expression in anterior pituitary cells as assessed by reverse transcriptase-polymerase chain reaction. The current results indicate that NO is involved in the inhibitory effect of TNF-α on prolactin secretion and that TNF-α induces iNOS transcription and stimulates NO synthesis in anterior pituitary cells.Fil: Theas, Maria Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: de Laurentiis, Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Candolfi, Marianela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: López, Silvia Liliana. Universidad de Buenos Aires. Facultad de Medicina; ArgentinaFil: Carrasco, Andrés E.. Universidad de Buenos Aires. Facultad de Medicina; ArgentinaFil: Zaldivar, Verónica. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Seilicovich, Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentin

Apoptosis of Lactotrophs Induced by D2 Receptor Activation Is Estrogen Dependent

BACKGROUND/AIMS: Dopamine (DA) inhibits prolactin release and reduces lactotroph proliferation by activating D2 receptors. DA and its metabolite, 6-hydroxydopamine (6-OHDA), induce apoptosis in different cell types. DA receptors and DA transporter (DAT) were implicated in this action. Considering that estradiol sensitizes anterior pituitary cells to proapoptotic stimuli, we investigated the effect of estradiol on the apoptotic action of DA and 6-OHDA in anterior pituitary cells, and the involvement of the D2 receptor and DAT in the proapoptotic effect of DA. METHODS: Viability of cultured anterior pituitary cells from ovariectomized rats was determined by MTS assay. Apoptosis was evaluated by Annexin-V/flow cytometry and TUNEL. Lactotrophs were identified by immunocytochemistry. RESULTS: DA induced apoptosis of lactotrophs in an estrogen-dependent manner. In contrast, estradiol was not required to trigger the apoptotic action of 6-OHDA. Cabergoline, a D2 receptor agonist, induced lactotroph apoptosis, while sulpiride, a D2 receptor antagonist, blocked DA-induced cell death. The blockade of DAT by GBR12909 did not affect the apoptotic action of DA, but inhibited 6-OHDA-induced apoptosis. CONCLUSION: These data show that DA, through D2 receptor activation, induces apoptosis of estrogen-sensitized anterior pituitary cells, and suggest that DA contributes to the control of lactotroph number not only by inhibiting proliferation but also by inducing apoptosisFil: Radl, Daniela Betiana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Zarate, Sandra Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Jaita, Gabriela Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Ferraris, Maria Jimena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Zaldivar, Verónica. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Eijo Alvarenga, Stella Guadalupe. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Seilicovich, Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; ArgentinaFil: Pisera, Daniel Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; Argentin

Estradiol increases the secretion and content of N-terminal PRL fragments from anterior pituitary cells in culture.

<p>Anterior pituitary cells from OVX rats were incubated in the presence of VEH or E2 for 24 h. A: Culture media and B: cells were obtained and processed for western blot analysis. Each column represents the mean ± SE of the relative increment of N-terminal PRL fragments with respect to VEH. In B, data of each column were normalized to β-actin expression (A, n = 6 wells/group; B, n = 6 wells/group), representative of three independent experiments. *p<0.05, **p<0.01 vs. VEH without E2, Student's t test. Lower panels: Representative blots of media (A) or cells (B) cultured with VEH or E2.</p

N-terminal PRL fragments content in the anterior pituitary varies along the estrous cycle.

<p>A: Recombinant 23 kDa PRL (r23-PRL), recombinant 16 kDa prolactin (r16-PRL) or pituitary protein extracts treated with β-mercaptoethanol (P+β, reducing conditions) or without β-mercaptoethanol (P, non-reducing conditions) were incubated with anti-recombinant rat PRL antibody (anti rrPRL, 1∶25000). The antibody recognizes both PRL forms. B: Anterior pituitaries from rats euthanized at diestrus I or proestrus were processed for western blot. Upper panel: Each column represents the mean ± SE of the relative increment of N-terminal PRL fragments content with respect to diestrus I. Data of each column were normalized to β-actin expression (n = 4–7 animals per group). *p<0.01 vs. diestrus I, Student's t test. Lower panel: Representative blot of pituitary proteins from rats euthanized at diestrus I or proestrus.</p

16 kDa PRL inhibits anterior pituitary cell proliferation.

<p>A and B: Anterior pituitary cells from OVX rats were incubated for 24 h with VEH and FSK, and with or without 16 kDa PRL (10 nM) for the last 4 h. D and E: Anterior pituitary cells from OVX rats were incubated for 24 h with E2, then for 1 h with E2 and FSK, and for the last 4 h in the same media with or without 16 kDa PRL (10 nM). Proliferation was determined by BrdU incorporation (3 h) and fluorescence microscopy. A and D: Each column represents the percentage ± CI (95%) of BrdU-positive cells (n≥1000 cells/group, left) or BrdU-positive lactotropes (n≥300 cells/group, right), representative of four independent experiments. Data were analyzed by χ². **p<0.01 vs. respective control without 16 kDa PRL. B and E: Representative images of anterior pituitary cells showing immunoreactivity for prolactin (red) counterstained with DAPI (blue) and BrdU (green). Arrowheads indicate proliferating lactotropes. Scale bar: 50 µm. C and F: Anterior pituitary cells in culture were incubated with VEH (C) or E2 (F) for 24 h, and with or without 16 kDa PRL (10 nM) for the last 4 h. Cell cycle was analyzed by flow cytometry using PI. Left: Each column represents the mean ± SE of the percentage of S-phase cells (n≥4 wells/group), representative of three independent experiments. Data were analyzed by Student's t test. *p<0.05 vs. respective control without 16 kDa PRL. Right: Representative histograms of DNA content of anterior pituitary cells incubated in the presence or absence of 16 kDa PRL.</p