Proangiogenic cytokines produced by non-Hodgkin lymphoma tumor cells induce angiogenesis in infiltrated bone marrow samples

Universidade Federal de São Paulo, Hematol & Transfus Serv, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Pathol, São Paulo, BrazilUniversidade Federal de São Paulo, Hematol & Transfus Serv, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Pathol, São Paulo, BrazilWeb of Scienc

Comparative Expression of a Set of Genes to an Internal Housekeeping Control in CDNA Amplified and not Amplified by PolyAPCR in Non-Hodgkin's Lymphoma Samples Obtained From Fine-Needle Aspiration Cytology

Aim: We aimed to evaluate the amount and quality of the RNA obtained from lymph nodes of non-Hodgkin lymphomas (NHLs) patients using fine-needle aspiration cytology (FNAC), and to develop strategies to overcome eventual technical drawbacks.Materials and Methods: Twenty-six patients with NHL and 10 tonsils from children submitted to tonsillectomy underwent FNAC. the aspirates were performed using both cytoaspirator (sample A) and syringe and needle (sample B). the RNA was extracted using Trizol reagent and transcribed with the Superscript kit (Invitrogen). the quality of RNA was verified through the amplification of a beta-actin 155-bp fragment.Results: Fifty-two NHL and 20 tonsil samples were analyzed. the total amount of RNA in the tonsil samples varied from <1.0 to 6.2 mu g with cytoaspirator (A) and from <1.0 to 4.7 mu g with syringe and needle (B). the total amount of RNA obtained from NHL varied from <1.0 to 6.5 mu g with cytoaspirator (A) and <1.0 to 5.5 mu g with syringe and needle. in an attempt to increase the amounts of RNA in each sample, we standardized the polyAPCR technique, which increased by 10 times the amount of cDNA in most of the test and control samples. the efficiency of the reaction was verified through the amplification of beta-actin, in which 100% of the test and control samples were amplified. When polyAPCR cDNA and nonamplified cDNA samples were paired to be evaluated by real-time PCR, using glyceraldehyde-3-phosphate dehydrogenase as the constitutive gene and nuclear factor-kappa B and NF kappa BIA as target genes, there was equivalence in the amplifications of 100% of the 15 evaluated samples.Conclusions: Our results showed that FNAC, obtained either by cytoaspirator or syringe and needle, is a good source of small amounts of RNA. the polyAPCR technique significantly increased the amount of genomic material, which might be a cDNA source for future gene expression studies.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Universidade Federal de São Paulo, EPM, Hematol & Hemotherapy Serv, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, EPM, Dept Pathol, BR-04023900 São Paulo, BrazilUNIFESP, EPM, Dept Biol Sci, Diadema, BrazilUniversidade Federal de São Paulo, EPM, Hematol & Hemotherapy Serv, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, EPM, Dept Pathol, BR-04023900 São Paulo, BrazilUNIFESP, EPM, Dept Biol Sci, Diadema, BrazilWeb of Scienc

Angiomirs expression profiling in diffuse large B-Cell lymphoma

Despite advances in treatment, 30% of diffuse large B-cell lymphoma (DLBCL) cases are refractory or relapse after chemoimmunotherapy. Currently, the relationship between angiogenesis and angiomiRs in DLBCL is unknown. We classified 84 DLBCL cases according to stromal signatures and evaluated the expression of pro-and antiangiomiRs in paraffin embedded tissues of DLBCL and correlated them with microvascular density (MVD). 40% of cases were classified as stromal-1, 50% as stromal-2 and 10% were not classified. We observed increased expression of proangiomiRs Let-7f, miR-17, miR-18a, miR-19b, miR-126, miR-130a, miR-210, miR-296 and miR-378 in 14%, 57%, 30%, 45%, 12%, 12%, 56%, 58% and 48% of the cases, respectively. Among antiangiomiRs we found decreased expression of miR-16, miR-20b, miR-92a, miR-221 and miR-328 in, respectively, 27%, 71%, 2%, 44% and 11%. We found association between increased expression of proangiomiRs miR-126 and miR-130a and antiangiomiR miR-328 and the subtype non-GCB. We found higher levels of the antiangiomiRs miR-16, miR-221 and miR-328 in patients with low MVD and stromal-1 signature. IPI and CD34 confirmed independent impact on survival of the study group. None of the above angiomiRs showed significance as biomarker in an independent serum samples cohort of patients and controls. In conclusion, we confirmed association between antiangiomiRs miR-16, miR-221 and miR-328 and stromal-1 signature. Four angiomiRs emerged as potential therapeutic targets: proangiomiRs miR-17, miR-210 and miR-296 and antiangiomiR miR-20b. Although the four microRNAs seem to be important in DLBCL pathogenesis, they were not predictive of DLBCL onset or relapse in the serum independent cohort.Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq), BrazilFundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP), BrazilUniv Fed Sao Paulo, Dept Oncol Clin & Expt, Sao Paulo, BrazilAC Camargo Canc Ctr, Sao Paulo, BrazilInsper Inst Educ & Res, Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Patol, Sao Paulo, BrazilUniv Texas MD Anderson Canc Ctr, Dept Hematopathol, Houston, TX 77030 USAUniv Fed Sao Paulo, Dept Oncol Clin & Expt, Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Patol, Sao Paulo, BrazilFAPESP: 2010/17668-6Web of Scienc