Rodrigo Calado | Universidade de São Paulo (original) (raw)

Papers by Rodrigo Calado

Haematologica, Jan 10, 2015

Graft-versus-host disease (GVHD) is a major risk factor for secondary malignancy after hematopoie... more Graft-versus-host disease (GVHD) is a major risk factor for secondary malignancy after hematopoietic stem cell transplantation. Squamous cell carcinoma (SCC) of the skin and mucous membranes are especially frequent in this setting where aneuploidy and tetraploidy are associated with aggressive disease. The current study is directed to the mechanism of neoplasia in this setting. Un-manipulated keratinocytes from areas of oral GVHD in 9 patients showed tetraploidy in 10-46% of cells when examined by florescence in situ hybridization (FISH). Keratinocytes isolated from biopsy sites of GVHD but not from normal tissue showed even greater numbers of tetraploid cells (mean 78%, range 15-85%; N=9) after culture. To mimic the inflammatory process in GVHD, allogeneic HLA-mismatched lymphocytes were mixed with normal keratinocytes. After two weeks, substantial numbers of aneuploid and tetraploid cells were evident in cultures with lymphocytes and with purified CD8 but not CD4 cells. Telomere length was substantially decreased in the lymphocyte-treated sample. No mutations were present in p53 gene, although haploinsufficiency for p53 due to loss of chromosome 17 was common in cells exposed to lymphocytes. These findings suggest that in GVHD, inflammation and repeated cell division correlates with the development of karyotypic abnormalities.

BMC medical genetics, 2014

Telomeres are repeated sequences (the hexanucleotide TTAGGG in vertebrates) located at chromosome... more Telomeres are repeated sequences (the hexanucleotide TTAGGG in vertebrates) located at chromosome ends of eukaryotes, protecting DNA from end joining or degradation. Telomeres become shorter with each cell cycle, but telomerase, a ribonucleoprotein complex, alleviates this attrition. The telomerase RNA component (TERC) is an essential element of telomerase, serving as a template for telomere elongation. The H/ACA domain of TERC is indispensable for telomere biogenesis. Mutations in the telomerase components allow accelerated telomere loss, resulting in various disease manifestations, including bone marrow failure. To date, this is the first detailed report of an H-box mutation in TERC that is related to human disease. A 26-year-old man with myelodysplastic syndrome (MDS) had very short telomeres. Sequencing identified a single heterozygous mutation in the H box of the patient's TERC gene. The same mutation was also present in his father and his son, demonstrating that it was ger...

POWERENG 2007 - International Conference on Power Engineering - Energy and Electrical Drives Proceedings, 2007

Power converter of a switched reluctance actuator can present different configurations, and sever... more Power converter of a switched reluctance actuator can present different configurations, and several control strategies can also be adopted, each one of them resulting in different actuator skills. For optimal actuator behaviour, and depending on target application, an appropriated control strategy must be developed and applied. The proposed methodology, here presented, uses the knowledge of static and dynamic actuator characterization

Hematology Journal, 2003

Introduction: Classically, the monocytic component of acute myelomonocytic and acute monocytic/mo... more Introduction: Classically, the monocytic component of acute myelomonocytic and acute monocytic/monoblastic (FAB-M5) leukemias is demonstrated by nonspecific esterase positivity in cytochemical stainings. We have previously demonstrated that nonspecific esterases from normal monocytes can be determined by a chemiluminescent method. In the present study, we investigated whether this assay can also determine the monocytic component of FAB-M4 and FAB-M5 and distinguish these acute myeloid leukemia (AML) categories. Materials and methods: Bone marrow samples were obtained from 66 patients with AML (M0, two cases; M1, 12 cases; M2, 13 cases; M3, 10 cases; M4, 11 cases; M5, 12 cases; M6, two cases; M7, four cases). Cells were incubated with a standard reaction mixture and chemiluminescence was measured for 10 min. Two parameters were assessed, the peak (PLE) and the integrated light emission (ILE). Results: Both PLE and ILE were higher in FAB-M4 and FAB-M5 subtypes compared to other AML subtypes (Po0.001). In addition, the classification of AML cases into FAB-M4, FAB-M5 and nonmonocytic subtypes based on ILE analysis was concordant with a-naphthyl acetate esterase (ANAE) in 97% of cases (kappa coefficient 0.94, Po0.001). Conclusions: These findings indicate that this chemiluminescent assay was able to determine the monocytic component of FAB-M4 and FAB-M5 cells, and the classification of AML subtypes based on chemiluminescent analysis strongly agreed with the cytochemical ANAE-staining. In conclusion, this chemiluminescent assay is a simple, fast and objective method, which may be useful as an alternative tool in the differential diagnosis of AML subtypes.

Cytometry, 1999

The presence of phenotypically immature lymphocytes in umbilical cord blood has been a controvers... more The presence of phenotypically immature lymphocytes in umbilical cord blood has been a controversial topic. Moreover, their changes with age have not been systematically evaluated. In the present study, relative and absolute numbers of CD34+, CD10+CD19+, and CD4+CD8+ cell subsets were determined in umbilical cord blood from 12 full-term normal newborns, 43 children aged 1 month to 6 years, and 10 young adults. The samples were processed by whole-blood lysis and monoclonal antibody staining, and cells were analyzed by flow cytometry. Immature cells were present in cord blood and progressively declined in both absolute and percentage numbers with age, each according to a particular curve, reaching youth values roughly at age 2-4 years. These results demonstrate that phenotypically immature cells normally circulate at low levels in peripheral blood, mostly at birth and during infancy, but also during youth.

International Journal of Cardiology, 2015

http://bloodjournal.hematologylibrary.org/content/104/6/1671.full.html Updated information and se... more http://bloodjournal.hematologylibrary.org/content/104/6/1671.full.html Updated information and services can be found at: (5019 articles) Immunobiology (3131 articles) Hematopoiesis and Stem Cells Articles on similar topics can be found in the following Blood collections http://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#repub_requests Information about reproducing this article in parts or in its entirety may be found online at: http://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#reprints Information about ordering reprints may be found online at: http://bloodjournal.hematologylibrary.org/site/subscriptions/index.xhtml Information about subscriptions and ASH membership may be found online at: Infusion of parental lymph node (LN) cells into sublethally irradiated hybrid F1 recipients created a murine model for bone marrow (BM) failure. Affected animals developed fatal pancytopenia within 2 to 3 weeks, accompanied by BM oligoclonal T-cell infiltration and severe marrow hypoplasia indicated by approximately 10fold declines in total BM cellularity, 15fold declines in BM Lin ؊ Sca1 ؉ c-Kit ؉ cells, 100-fold declines in spleen colony-forming units, and 100-fold declines in hematopoietic progenitor and stem cells as esti-mated by irradiation protection in vivo. LN cells of both H2 b/b and H2 d/d haplotypes were effectors. Serum interferon-␥ (IFN-␥) concentration increased 2-to 3-fold. Marrow cells were severely apoptotic, with high proportions of Fas ؉ and annexin V ؉ cells. Cotransplantation of 5 ؋ 10 5 BM cells from clinically affected donors and 10 6 BM cells from H2 identical healthy mice could not rescue lethally irradiated recipients. Recipients had significantly lower cellularity in peripheral blood and BM, and cell mixtures failed to produce a stromal feeder layer to support marrow cell growth in vitro. Pathogenic T cells from donors after BM failure appeared capable of destroying hematopoietic progenitor, stem, and stromal cells from fully compatible healthy donors as "innocent bystanders." This effect can be partially abrogated by anti-IFN-␥ antibody.

Experimental Hematology, 2005

Materials and Methods. Sublethally irradiated CByB6F1 mice were infused with 5 × 10 6 C57BL/ 6 (B... more Materials and Methods. Sublethally irradiated CByB6F1 mice were infused with 5 × 10 6 C57BL/ 6 (B6) lymph node (LN) cells. Recipient BM cells were taken at 3, 7, 10, and 14 days following LN infusion and were cultured in vitro in a-modified Eagle media for 2-3 weeks. Peripheral blood and was analyzed by complete blood counts while BM lymphocyte infiltration/expansion was analyzed by flow cytometry. Marrow cells from affected and control mice were mixed and cultured in vitro to test nonspecific stromal damage. Results. Donor lymphocytes infiltrated host BM within 3-7 days and expanded significantly between 7 and 10 days, concurrent with the development of leukopenia, thrombocytopenia, and marrow hypoplasia. BM cells from mice at 7, 10, and 14 days after B6-LN cell infusion were progressively defective in forming stromal feeder layers. A 1:1 mixture of BM cells from affected CByB6F1 mice and normal B6 mice failed to form an effective stromal feeder layer that could support cobblestone colony formation, indicating that lymphocytes in the BM of affected CByB6F1 mice were able to damage stromal cells in the normal B6 BM.

Brazilian Journal of Medical and Biological Research, 2000

Although iron can catalyze the production of free radicals involved in LDL lipid peroxidation, th... more Although iron can catalyze the production of free radicals involved in LDL lipid peroxidation, the contribution of iron overload to atherosclerosis remains controversial. The description of two mutations in the HFE gene (Cys282Tyr and His63Asp) related to hereditary hemochromatosis provides an opportunity to address the question of the association between iron overload and atherosclerosis. We investigated the prevalence of HFE mutations in 160 survivors of myocardial infarction with angiographically demonstrated severe coronary atherosclerotic disease, and in 160 age-, gender-and race-matched healthy control subjects. PCR amplification of genomic DNA followed by RsaI and BclI restriction enzyme digestion was used to determine the genotypes. The frequency of the mutant Cys282Tyr allele was identical among patients and controls (0.022; carrier frequency, 4.4%), whereas the mutant His63Asp allele had a frequency of 0.143 (carrier frequency, 27.5%) in controls and of 0.134 (carrier frequency, 24.5%) in patients. Compound heterozygotes were found in 2 of 160 (1.2%) controls and in 1 of 160 (0.6%) patients. The finding of a similar prevalence of Cys282Tyr and His63Asp mutations in the HFE gene among controls and patients with coronary atherothrombotic disease, indirectly questions the possibility of an association between hereditary hemochromatosis and atherosclerosis.

PLoS ONE, 2014

Background: Although implicated in the pathogenesis of several chronic inflammatory disorders and... more Background: Although implicated in the pathogenesis of several chronic inflammatory disorders and hematologic malignancies, telomerase mutations have not been thoroughly characterized in human cancers. The present study was performed to examine the frequency and potential clinical relevance of telomerase mutations in esophageal carcinomas.

Blood, Jan 22, 2015

The pathophysiology of severe aplastic anemia (SAA) is immune-mediated destruction of hematopoiet... more The pathophysiology of severe aplastic anemia (SAA) is immune-mediated destruction of hematopoietic stem and progenitor cells (HSPCs). Most patients respond to immunosuppressive therapies, but a minority transform to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML), frequently associated with monosomy 7 (-7). Thirteen SAA patients were analyzed for acquired mutations in myeloid cells at the time of evolution to -7, and all had a dominant HSPC clone bearing specific acquired mutations. However, mutations in genes associated with MDS/AML were present in only 4 cases. Patients who evolved to MDS and AML showed marked progressive telomere attrition before the emergence of -7. Single telomere length analysis confirmed accumulation of short telomere fragments of individual chromosomes. Our results indicate that accelerated telomere attrition in the setting of a decreased HSPC pool is characteristic of early myeloid oncogenesis, specifically chromosome 7 loss, in MDS/AML aft...

F1000 Medicine Reports, 2012

Telomeres and telomere repair are basic molecular features of cells possessing linear DNA chromos... more Telomeres and telomere repair are basic molecular features of cells possessing linear DNA chromosomes and defects in them result in various diseases. This review examines recent advances in understanding these diseases, particularly at a molecular level, and in relating telomere dysfunction to clinical diseases. We also discuss the potential role of telomere elongation as a therapy in diseases, and more controversially, the prevention/reversal of aging.

Fertility and Sterility, 2014

Objective: To analyze whether leukocyte telomere length (LTL) is impaired in women with polycysti... more Objective: To analyze whether leukocyte telomere length (LTL) is impaired in women with polycystic ovary syndrome (PCOS). Design: Case-control study. Setting: Hospital. Patient(s): A total of 274 women, including 150 patients with PCOS and 124 controls. Intervention(s): None. Main Outcome Measure(s): Body mass index (BMI), waist circumference, systemic arterial pressure, lipid profile, E 2 , LH, T, androstenedione, PRL, TSH, sex hormone-binding globulin, C-reactive protein (CRP), homocysteine, free androgen index, and the homeostatic model of insulin sensitivity (HOMA-IR) index were analyzed. The LTL evaluation was measured by quantitative polymerase chain reaction. Result(s): The PCOS group had higher values for weight, BMI, waist circumference, systolic arterial pressure, triglycerides, LH, T, insulin, CRP, free androgen index, and HOMA-IR compared with the control group. Sex hormone-binding globulin and E 2 levels were lower in the PCOS group than in the control group. The LTL did not differ between groups. Age, BMI, and HOMA-IR had no significant effect on LTL. The inflammatory biomarkers CRP and homocysteine were negatively correlated with LTL in patients with PCOS. Conclusion(s): Our results showed no differences in LTL between patients with PCOS and controls, but CRP and homocysteine biomarkers negatively correlated with LTL in the PCOS group. (Fertil Steril Ò 2015;103:542-7. Ó2015 by American Society for Reproductive Medicine.)

Experimental Hematology, 2006

Objective. Abnormal telomere shortening has been observed in a subset of individuals with aplasti... more Objective. Abnormal telomere shortening has been observed in a subset of individuals with aplastic anemia (AA). We hypothesized that genetic variation in two genes critical in telomere biology, TERF1 and TERF2, could be a risk factor for AA. Methods. The proximal promoter and all coding regions of TERF1 and TERF2 were sequenced in 47 individuals with acquired AA. Regions with genetic variation were sequenced in an additional 95 AA patients and 289 healthy controls. Single nucleotide polymorphism (SNP) frequencies were analyzed using co-dominant and dominant models and haplotypes determined. Functional studies evaluated telomerase activity, telomere and telomeric overhang lengths, and TRF2 protein expression in select patients. Results. Two nonsynonymous amino acid changes were detected, one in exon 9 of TERF1 and another in exon 6 of TERF2. These sequence variants resulted in conservative amino acid changes and were not predicted to alter TRF1 or TRF2 protein expression or function. SNP and haplotype analyses in acquired AA patients suggested that one variant allele, in intron 9 of TERF1, and haplotype could be associated with increased risk for aplastic anemia (OR 1.59, 95% confidence interval 1.06-2.39, p [ 0.033). TERF2 SNPs and haplotypes were not significantly associated with aplastic anemia. Conclusions. It is possible that a common genetic variant in TERF1 is associated with risk for AA but additional studies are required. Highly penetrant, non-synonymous, or insertiondeletion mutations in TERF1 and TERF2 were not identified and therefore are not likely to be major genetic risk factors for the development of AA. Ó

PLoS ONE, 2012

We previously reported the development of a lethal myeloid sarcoma in a non-human primate model u... more We previously reported the development of a lethal myeloid sarcoma in a non-human primate model utilizing retroviral vectors to genetically modify hematopoietic stem and progenitor cells. This leukemia was characterized by insertion of the vector provirus into the BCL2A1 gene, with resultant BCL2A1 over-expression. There is little information on the role of this anti-apoptotic member of the BCL2 family in hematopoiesis or leukemia induction. Therefore we studied the impact of Bcl2a1a lentiviral over-expression on murine hematopoietic stem and progenitor cells. We demonstrated the anti-apoptotic function of this protein in hematopoietic cells, but did not detect any impact of Bcl2a1a on in vitro cell growth or cell cycle kinetics. In vivo, we showed a higher propensity of HSCs over-expressing Bcl2a1a to engraft and contribute to hematopoiesis. Mice over-expressing Bcl2a1a in the hematologic compartment eventually developed an aggressive malignant disease characterized as a leukemia/lymphoma of B-cell origin. Secondary transplants carried out to investigate the primitive origin of the disease revealed the leukemia was transplantable. Thus, Bcl2a1 should be considered as a protooncogene with a potential role in both lymphoid and myeloid leukemogenesis, and a concerning site for insertional activation by integrating retroviral vectors utilized in hematopoietic stem cell gene therapy.

New England Journal of Medicine, 2005

Mutations in TERC, the gene for the RNA component of telomerase, cause short telomeres in congeni... more Mutations in TERC, the gene for the RNA component of telomerase, cause short telomeres in congenital aplastic anemia and in some cases of apparently acquired hematopoietic failure. We investigated whether mutations in genes for other components of telomerase also occur in aplastic anemia.