Η μη επαγόμενη και η επαγόμενη παραγωγή των GM-CSF, G-CSF, IL-3, IL-6, SCF, IL-1β, TNF-α , TNF-β, IFN-γ και του TGF-β προσδιορίστηκαν μετά από καλλιέργεια μονοπύρηνων κυττάρων αίματος 22 ασθενών με βαριά β-θαλασσαιμία που υποβάλλονταν σε κανονικό πρόγραμμα μεταγγίσεων, 5 ασθενών με ενδιάμεση β- θαλασσαιμία που δεν υποβάλλονταν σε κανονικό πρόγραμμα μεταγγίσεων και 9 φυσιολογικών ατόμων. Ένα διακριτό πρότυπο παραγωγής κυτταροκινών διαπιστώθηκε στους β-θαλασσαιμικούς ασθενείς. Συγκεκριμένα παρατηρήθηκε μειωμένη μη επαγόμενη παραγωγή όλων των κυτταροκινών και σημαντική αύξηση της επαγόμενης παραγωγής των IFN-γ, TNF-α και της IL-1β. Αυτές οι διαταραχές ήταν περισσότερο εμφανείς στους μεγαλύτερους σε ηλικία ασθενείς, που έχουν υποβληθεί στις περισσότερες μεταγγίσεις. Η αυξημένη παραγωγή των παραπάνω κυτταροκινών, συνήθως χαρακτηρίζει την οξεία απόκριση σε μολυσματικούς παράγοντες και ασκεί αρνητική επίδραση στην ερυθροποίηση. Αυτό μπορεί να εξηγεί την επιδείνωση της αναιμίας στους θαλασσαιμικούς ασθενείς, κατά τη διάρκεια οξείων λοιμώξεων. / The unstimulated and induced production of granulocyte-macrofage colonystimulating factor (GM-CSF), granulocyte colony –stimulating factor (G-CSF), IL-3, IL-6, stem cell factor (SCF), IL-1β, tumour necrosis factor-alpha (TNF-a), TNF-β, interferon-gamma (IFN-γ) and transforming growth factor-beta (TGF-β) was determined after culture of blood mononuclear cells from 22 patients with severe β- thalassaemia in a regular transfusion programme, five non-regularly transfused patients with β-thalassaemia intermedia and nine normal persons. A distinct pattern of cytokine production in thalassaemic patients was detected, namely a low unstimulated production of all cytokines and a significant increase in the stimulated production of IFN-γ, TNF-a and IL-1β; these abnormalities were more pronounced in the more heavily transfused older patients. The increased production of the above cytokines, which usually characterize the acute response to infectious agents and have a negative effect on erythropoiesis, may explain the deterioration of anaemia found in thalassaemic patients during acute infections.

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Objetivos: Avaliar o potencial terapêutico e a segurança do uso intravítreo de (FMMO) contendo células CD34+ em pacientes portadores de degeneração macular relacionada com a idade na forma atrófica. Casuística e Métodos: Foram avaliados 10 pacientes com degeneração macular relacionada à idade (DMRI) seca e acuidade visual no pior olho <=20/100. Foi obtido aspirado da medula óssea de todos os pacientes, e após o processamento do material no hemocentro, foi injetado 0,1 ml da suspensão de FMMO intravítreo no olho com pior acuidade. Os pacientes foram avaliados no baseline, 1, 3, 6, 9 e 12 meses após a injeção. Todos realizaram medida da melhor acuidade visual corrigida (MAVC), microperimetria, eletrorretinografia multifocal (ERGmf), autofluorescência, Tomografia de coerência óptica (OCT) e responderam o questionário VFQ-25 em todos seguimentos. Também foi realizada angiografia fluoresceínica antes da injeção, seis e doze meses após. Resultados: Todos os pacientes completaram o seguimento de seis meses, e seis finalizaram os doze meses. Antes da injeção, a média da MAVC foi de 1,18 logMAR (20/320-1); variando de 20/125 a 20/640-2. Aos doze meses, a média foi de 1,0 logMAR (20/200), com melhora significativa em todos os meses de seguimento. A média do limiar de sensibilidade na microperimetria mostrou melhora significativa a partir do sexto mês (p=0,009). Ao se dividirem os pacientes com área de atrofia maior e menor, por meio da medida da hipoautofluorescência, observou-se que a melhora foi significativa apenas no grupo de menor atrofia. Não houve diferença significativa na ERGmf. A angiofluoresceinografia não apresentou crescimento de neovasos ou tumores. O questionário de qualidade de vida mostrou diferença significativa na saúde mental (p=0,003) e na visão de cores (p=0,005) e forte tendência à significância na análise que abordou a visão geral (p=0,05) e dependência (p=0,067). Houve declínio em relação à saúde geral (p=0,77). Conclusões: Os resultados indicam que o uso de FMMO intravítreo em pacientes com DMRI é seguro e está associado à melhora da acuidade visual e microperimetria. Pacientes com área menor de atrofia têm melhor resposta. Não é possível afirmar, mas acredita-se no resgate funcional das células em sofrimento, que ainda não se degeneraram. / Objectives: To assess the therapeutic potential and safety of the intraviteral use of a bone marrow mononuclear fraction (BMMF) containing CD 34+ cells in patients with the atrophic form of age-related macular degeneration (AMD). Casuistic and Methods: The study was conducted on 10 patients with the atrophic form of ARMD with worse eye visual acuity <=20/100. Bone marrow was aspirated from each patient under local anesthesia. After processing and separation of mononuclear cells, 0.1 ml of the suspension was injected intravitreally into the eye of lower acuity. The patients were evaluated at baseline and 1, 3, 6, 9 and 12 months after the injection. All were submitted to measurement of best corrected visual acuity (BCVA), microperimetry, multifocal electroretinography (ERGmf), autofluorescence, and optical coherence tomography (OCT) and all responded to the VFQ-25 questionnaire at all follow-up visits. Fluorescein angiography was performed before the injection and six and 12 months later. Results: All patients completed the six month follow-up and six completed the 12 month follow-up after the injection. Before the injection, mean BCVA was 1.18 logMAR (20/320-1), ranging from 20/125 to 20/640-2 and at 12 months it was 1.0 logMAR (20/200), with a significant improvement over all followup months. Mean perimetric threshold sensitivity improved significantly starting at the sixth month (p=0.009). When the patients with a larger area of atrophy measured by hypoautofluorescence were considered separately, a significant improvement was observed only in the group with lower atrophy. There was no significant difference in electroretinography. Angiofluoresceinography did not reveal neovessel or tumor growth. The quality of life questionnaire showed a significant difference in mental health (p=0.003) and color vision (p=0.005), and a strong tendency in the analysis of general vision (p=0.05) and dependence (p=0.067). There was a decline in general health (p=0.77). Conclusions: The results indicate that the use of intravitreal BMMF in patients with AMD is safe and is associated with improved visual acuity and microperimetry. Patients with a smaller atrophy area obtained a better response. The paracrine effect of these cells may explain the functional improvement observed in the present study; however, a larger series should be study to confirm these clinical findings.

Objetivos: Avaliar o potencial terapêutico e a segurança do uso intravítreo de (FMMO) contendo células CD34+ em pacientes portadores de degeneração macular relacionada com a idade na forma atrófica. Casuística e Métodos: Foram avaliados 10 pacientes com degeneração macular relacionada à idade (DMRI) seca e acuidade visual no pior olho <=20/100. Foi obtido aspirado da medula óssea de todos os pacientes, e após o processamento do material no hemocentro, foi injetado 0,1 ml da suspensão de FMMO intravítreo no olho com pior acuidade. Os pacientes foram avaliados no baseline, 1, 3, 6, 9 e 12 meses após a injeção. Todos realizaram medida da melhor acuidade visual corrigida (MAVC), microperimetria, eletrorretinografia multifocal (ERGmf), autofluorescência, Tomografia de coerência óptica (OCT) e responderam o questionário VFQ-25 em todos seguimentos. Também foi realizada angiografia fluoresceínica antes da injeção, seis e doze meses após. Resultados: Todos os pacientes completaram o seguimento de seis meses, e seis finalizaram os doze meses. Antes da injeção, a média da MAVC foi de 1,18 logMAR (20/320-1); variando de 20/125 a 20/640-2. Aos doze meses, a média foi de 1,0 logMAR (20/200), com melhora significativa em todos os meses de seguimento. A média do limiar de sensibilidade na microperimetria mostrou melhora significativa a partir do sexto mês (p=0,009). Ao se dividirem os pacientes com área de atrofia maior e menor, por meio da medida da hipoautofluorescência, observou-se que a melhora foi significativa apenas no grupo de menor atrofia. Não houve diferença significativa na ERGmf. A angiofluoresceinografia não apresentou crescimento de neovasos ou tumores. O questionário de qualidade de vida mostrou diferença significativa na saúde mental (p=0,003) e na visão de cores (p=0,005) e forte tendência à significância na análise que abordou a visão geral (p=0,05) e dependência (p=0,067). Houve declínio em relação à saúde geral (p=0,77). Conclusões: Os resultados indicam que o uso de FMMO intravítreo em pacientes com DMRI é seguro e está associado à melhora da acuidade visual e microperimetria. Pacientes com área menor de atrofia têm melhor resposta. Não é possível afirmar, mas acredita-se no resgate funcional das células em sofrimento, que ainda não se degeneraram. / Objectives: To assess the therapeutic potential and safety of the intraviteral use of a bone marrow mononuclear fraction (BMMF) containing CD 34+ cells in patients with the atrophic form of age-related macular degeneration (AMD). Casuistic and Methods: The study was conducted on 10 patients with the atrophic form of ARMD with worse eye visual acuity <=20/100. Bone marrow was aspirated from each patient under local anesthesia. After processing and separation of mononuclear cells, 0.1 ml of the suspension was injected intravitreally into the eye of lower acuity. The patients were evaluated at baseline and 1, 3, 6, 9 and 12 months after the injection. All were submitted to measurement of best corrected visual acuity (BCVA), microperimetry, multifocal electroretinography (ERGmf), autofluorescence, and optical coherence tomography (OCT) and all responded to the VFQ-25 questionnaire at all follow-up visits. Fluorescein angiography was performed before the injection and six and 12 months later. Results: All patients completed the six month follow-up and six completed the 12 month follow-up after the injection. Before the injection, mean BCVA was 1.18 logMAR (20/320-1), ranging from 20/125 to 20/640-2 and at 12 months it was 1.0 logMAR (20/200), with a significant improvement over all followup months. Mean perimetric threshold sensitivity improved significantly starting at the sixth month (p=0.009). When the patients with a larger area of atrophy measured by hypoautofluorescence were considered separately, a significant improvement was observed only in the group with lower atrophy. There was no significant difference in electroretinography. Angiofluoresceinography did not reveal neovessel or tumor growth. The quality of life questionnaire showed a significant difference in mental health (p=0.003) and color vision (p=0.005), and a strong tendency in the analysis of general vision (p=0.05) and dependence (p=0.067). There was a decline in general health (p=0.77). Conclusions: The results indicate that the use of intravitreal BMMF in patients with AMD is safe and is associated with improved visual acuity and microperimetry. Patients with a smaller atrophy area obtained a better response. The paracrine effect of these cells may explain the functional improvement observed in the present study; however, a larger series should be study to confirm these clinical findings.

Apoptosis is a form of programmed cell death that is essential in such processes as organ and tissue remodeling and maturation of hematopoietic cells. The clearance of apoptotic cells is essential to prevent autoimmune responses to sequestered antigens. This process is mediated by phagocytes of the monocyte lineage. Before phagocytosis can occur, macrophages must be recruited to the apoptotic cells through chemotaxis. Products of the reaction catalyzed by the phospholipases A2 (PLA2) have been shown to induce monocyte chemotaxis either directly or indirectly. Some investigators have implicated a cytosolic calcium-independent PLA2 (iPLA2) in the production of these products during apoptosis. However, a recent report suggests that the secreted group IIa (sPLA2) binds to surfaces of apoptotic cells. The "receptor" for this pool of sPLA2 is the rod domain of vimentin, an intermediate filament protein that is exposed by caspase activity when cells undergo apoptosis. Based on these observations, we hypothesize that the exposure of vimentin on apoptotic cells traps a pool of catalytically active sPLA2 that then generates the bioactive lipids that induce macrophage chemotaxis. In our methods, [3H]-oleate labeled E-coli is used as a substrate for sPLA2 and enzyme activity is quantified by scintillation counting of released radiolabeled oleic acid. Apoptosis is induced with anti-fas (CD-95) on Jurkat cells and monitored through annexin-V binding and propidium iodide(PI) staining followed by flow cytometric analyses. THP-1 monocytes are employed in chemotaxis assay with monocyte chemotactic protein (MCP-1) as a positive control. The preliminary data show equivalent group IIa sPLA2 association with anti-fas treated and control cells, and the enzyme remains catalytically active when bound. In line with the hypothesis, trapped sPLA2 generated soluble molecules that directly or indirectly induced migration of THP-1 monocytes. However, the similar binding effect observed with apoptotic or control cells is surprising and experiments are being planned to determine if the interaction between IIa sPLA2 and apoptotic cells is vimentin dependent.

Multipotent hematopoietic stem cells (HSCs) are progenitors of all types of hematopoietic cells, and the efficient isolation and propagation of HSCs will significantly enhance our ability to manage many human disorders with bone marrow transplantation, stem cell transplantation and gene therapy. We employed "Signal Sequence Trap (SST)" method with yeast invertase to clone proteins on the surface of or secreted by stromal cells that enhance or inhibit the propagation of HSC’s in culture. AFT024, a mouse fetal liver stromal cell line that maintains stem cell activity in long-term culture, was subjected to SST analysis. We identified more than 60 signal sequences or transmembrane domain containing genes expressed by AFT024 cells. We compared their expression levels between AFT024 cells and BFC012 cells, a mouse fetal liver stromal cell line that was developed in the same way as for AFT024 cells but could not support HSC in long-term culture. Pleiotrophin, T16, Sca-1, deltalike and cytokine receptor like-1(CLF-1) are expressed significantly higher in AFT024 cells than in BFC012 cells. We recently employed Affymatrix genechip technology to study the interaction of HSCs and their microenvironment. In genechip experiments, Sca-1, deltalike, pleiotrophin and CLF-1 are among the most differentially expressed genes between AFT024 and BFC012 cells, while T16 was not represented on the chip. In addition, osteopontin, pigment epithelium-derived factor, proliferins, activin subunit, CXC chemokines GRO1 and LIX are more abundant in AFT024 cells than in BFC012 cells. Genechip technology was also applied to bone marrow stromal cell lines, including MS5, S17 and OP9 cells. Two murine multipotent hematopoietic cell lines, FDCP.mix and EML cells, were also analyzed. Data from these experiments are presented. / Singapore-MIT Alliance (SMA)

The receptor tyrosine kinase (RTK) c-Kit is expressed in hematopoietic stem and progenitor cells, mast cells and in several non-hematopoietic tissues. In the hematopoietic system c-Kit and its ligand Steel Factor (SF, aka Stem Cell Factor) are critical for proliferation, survival and differentiation. Mutations in either receptor or ligand lead to lethal anaemia, hematopoietic stem cell defects, mast cell deficiency and a series of non-hematological defects. The aims of the studies included in this thesis are to describe the signalling pathways downstream c-Kit in hematopoietic stem/progenitor cells and to further analyse the role of c-Kit signalling in fundamental biological functions. To study c-Kit signalling in the hematopoietic system we have employed hematopoietic stem cell-like cell lines which share many properties with primary hematopoietic stem cells in vitro and in vivo, including surface markers, multipotentiality, capacity for self-renewal and long term repopulation. In paper I we demonstrate that upon SF activation the RTK c-Kit is autophosphorylated and downstream signalling mediators are transiently activated. Surprisingly we find that the c-Kit mediated activation of the MAPK pathway is dependent on the activation of phosphoinositide 3-kinase (PI3K) in hematopoietic progenitor cells and that differentiation of these progenitors to mast cells results in a signalling switch where Raf activation changes from PI3K dependent to PI3K independent. We here establish that PI3K activity is required for viability and proliferation of hematopoietic progenitor cells. In paper II we studied the conventional protein kinase C (cPKC) involvement in c-Kit signalling. We observe that the cPKCs can phosphorylate c-Kit on serine 746 and that this phosphorylation negatively regulates the activation of the receptor. We demonstrate that inhibition of this negative phosphorylation results in dramatically increased protein kinase B (PKB) activation and as a consequence inhibition of cPKCs rescues cells from starvation induced apoptosis. Moreover we exhibit that the cPKCs are necessary for full activation of extracellular signal-regulated kinase (Erk) and that impaired PKC activity leads to hampered proliferation. In paper III we demonstrate that in addition to the cPKCs also the novel PKC is required for Erk activation and proliferation. Furthermore we present results indicating that PKC negatively regulates differentiation of bone marrow. In conclusion, with the studies in this thesis we display details in the signalling pathways induced upon RTK c-Kit activation and we demonstrate that c-Kit has significant effects on hematopoietic cell-physiology.

A síndrome mielodisplásica (SMD) é uma doença hematológica heterogênea, caracterizada por hematopoese anormal, displasia e instabilidade genômica, portanto, a análise citogenética é determinante no diagnóstico, prognóstico e acompanhamento evolutivo da doença. Considerando que as células hematopoéticas (CHs) e as estromais mesenquimais multipotentes (CTMs) estão em estreita associação, estudos que visem à caracterização destas poderão contribuir para elucidar os mecanismos que governam a progressão tumoral e identificar novos alvos terapêuticos. Objetivo: Caracterizar e comparar as CHs e CTMs derivadas de pacientes através da citogenética convencional e molecular para os genes aurora cinase A e B. Avaliar as propriedades biológicas das CTMs derivadas de SMD e controles saudáveis. Métodos: o estudo iniciou-se com a avaliação clinica de 25 pacientes e 8 controles saudáveis doo HCFMRP-USP e HAC-Jaú. Em seguida, foi realizada a análise cariótipica das CHs e CTMs da medula óssea pelo bandamento G e por FISH para os genes aurora A e B e o perfil imunofenotípico, bem como potencial de diferenciação em adipócito e osteócito das CTMs de pacientes portadores de SMD e controles saudáveis. Resultados: A avaliação clínica mostrou plaquetopenia (76%), neutropenia (100%), hemoglobina baixa (16%). A análise citogenética das CHs revelou cariótipo alterado em 13 pacientes (52%), com cariótipo complexo resultando em alterações numéricas e estruturais. Ao contrário, nas CTMs, o cariótipo se mostrou alterado em sete pacientes (28%) e um padrão de menor complexidade, apenas quatro pacientes apresentaram alterações nas duas populações celulares, porém, diferentes. Foram encontradas apenas alterações numéricas (sendo 86% monossomia e 14% ganho de cromossomo). As CHs e CTMs dos controles apresentaram cariótipos 100% normais. Na análise de FISH não foi evidenciada amplificação dos genes AURKA e AURKB. As CTMs dos pacientes e controles apresentaram-se semelhantes quanto à morfologia e potencial de diferenciação. Entretanto, as CTMs de pacientes mostraram-se alteradas para dois antígenos de superfície, CD90 e CD146, os quais mostraram níveis de expressão mais elevados nas amostras dos pacientes (p= 0,04, p = 0,001 respectivamente). Conclusão: Observou-se que as CTMs se encontram alteradas embora em menor frequência e diferindo das alterações encontradas nas CHs. Esses dados sugerem que as CTMs devem exercer importante papel na progressão tumoral e devem ser consideradas como alvos na busca de novas terapias e melhor esclarecimento dos mecanismos que governam a progressão tumoral. Apesar de não ter evidenciado amplificação dos genes AURKA e AURKB em SMD, estudos futuros que visem avaliar o nível de expressão dessas enzimas em pacientes portadores ou não de alterações citogenéticas poderão contribuir para a compreensão do envolvimento ou não desse gene com a evolução da doença. Além disso, não foi evidenciada associação de anemia profunda e citogenética alterada. / The myelodysplastic syndrome (MDS) is a heterogeneous hematologic disease characterized by abnormal hematopoiesis, dysplasia and genomic instability, therefore, cytogenetic analysis is crucial in the diagnosis, prognosis and monitoring of disease evolution. Whereas hematopoietic cells (CHs) and stromal multipotent mesenchymal (MSCs) are in close association studies aimed at the characterization of these may help to elucidate the mechanisms that govern tumor progression and identify novel therapeutic targets. Objective: To characterize and compare the CHs and MSCs derived from patients by conventional cytogenetics and molecular genes aurora kinase A and B. To evaluate the biological properties of MSCs derived from MDS and healthy controls. Methods: The study began with the clinical evaluation of 25 patients and eight healthy controls HCFMRP dooUSP and CH-Jau. Next, we performed a karyotypic analysis of CHs and MSCs from bone marrow by G-banding and FISH for aurora A and B genes and immunophenotypic profile and potential to differentiate into adipocytes and osteocytes of MSCs in patients with MDS and controls healthy. Results: The clinical evaluation showed thrombocytopenia (76%), neutropenia (100%), low hemoglobin (16%). The cytogenetic analysis revealed karyotype of CHs changed in 13 patients (52%), resulting in complex karyotype with numerical and structural changes. In contrast, in MSC, the karyotype was abnormal in seven patients (28%) and a pattern of lower complexity, only four patients had changes in both cell populations, however, different. Were found only numerical changes (monosomy being 86% and 14% gain in chromosome). The CHs and MSCs controls showed 100% normal karyotypes. In FISH analysis there was no evidence of gene amplification and AURKA AURKB. The MSCs of patients and controls were similar regarding the morphology and differentiation potential. However, the CTMs of patients proved to be changed to two surface antigens, CD90 and CD146, which showed higher expression levels in samples of patients (p = 0.04, p = 0.001 respectively). Conclusion: Furthermore, it was observed that the MSCs are changed although less frequently and differing from changes found in CHs. These data suggest that MSCs should play an important role in tumor progression and should be considered as targets in the search for new therapies and better explain the mechanisms that govern tumor progression. Although not shown AURKA amplification of genes in MDS and AURKB, future studies aimed at assessing the level of expression of these enzymes in patients with or without cytogenetic alterations may contribute to the understanding of the involvement or not of this gene with the disease. This study can not associate with profound anemia cytogenetic changes.

A síndrome mielodisplásica (SMD) é uma doença hematológica heterogênea, caracterizada por hematopoese anormal, displasia e instabilidade genômica, portanto, a análise citogenética é determinante no diagnóstico, prognóstico e acompanhamento evolutivo da doença. Considerando que as células hematopoéticas (CHs) e as estromais mesenquimais multipotentes (CTMs) estão em estreita associação, estudos que visem à caracterização destas poderão contribuir para elucidar os mecanismos que governam a progressão tumoral e identificar novos alvos terapêuticos. Objetivo: Caracterizar e comparar as CHs e CTMs derivadas de pacientes através da citogenética convencional e molecular para os genes aurora cinase A e B. Avaliar as propriedades biológicas das CTMs derivadas de SMD e controles saudáveis. Métodos: o estudo iniciou-se com a avaliação clinica de 25 pacientes e 8 controles saudáveis doo HCFMRP-USP e HAC-Jaú. Em seguida, foi realizada a análise cariótipica das CHs e CTMs da medula óssea pelo bandamento G e por FISH para os genes aurora A e B e o perfil imunofenotípico, bem como potencial de diferenciação em adipócito e osteócito das CTMs de pacientes portadores de SMD e controles saudáveis. Resultados: A avaliação clínica mostrou plaquetopenia (76%), neutropenia (100%), hemoglobina baixa (16%). A análise citogenética das CHs revelou cariótipo alterado em 13 pacientes (52%), com cariótipo complexo resultando em alterações numéricas e estruturais. Ao contrário, nas CTMs, o cariótipo se mostrou alterado em sete pacientes (28%) e um padrão de menor complexidade, apenas quatro pacientes apresentaram alterações nas duas populações celulares, porém, diferentes. Foram encontradas apenas alterações numéricas (sendo 86% monossomia e 14% ganho de cromossomo). As CHs e CTMs dos controles apresentaram cariótipos 100% normais. Na análise de FISH não foi evidenciada amplificação dos genes AURKA e AURKB. As CTMs dos pacientes e controles apresentaram-se semelhantes quanto à morfologia e potencial de diferenciação. Entretanto, as CTMs de pacientes mostraram-se alteradas para dois antígenos de superfície, CD90 e CD146, os quais mostraram níveis de expressão mais elevados nas amostras dos pacientes (p= 0,04, p = 0,001 respectivamente). Conclusão: Observou-se que as CTMs se encontram alteradas embora em menor frequência e diferindo das alterações encontradas nas CHs. Esses dados sugerem que as CTMs devem exercer importante papel na progressão tumoral e devem ser consideradas como alvos na busca de novas terapias e melhor esclarecimento dos mecanismos que governam a progressão tumoral. Apesar de não ter evidenciado amplificação dos genes AURKA e AURKB em SMD, estudos futuros que visem avaliar o nível de expressão dessas enzimas em pacientes portadores ou não de alterações citogenéticas poderão contribuir para a compreensão do envolvimento ou não desse gene com a evolução da doença. Além disso, não foi evidenciada associação de anemia profunda e citogenética alterada. / The myelodysplastic syndrome (MDS) is a heterogeneous hematologic disease characterized by abnormal hematopoiesis, dysplasia and genomic instability, therefore, cytogenetic analysis is crucial in the diagnosis, prognosis and monitoring of disease evolution. Whereas hematopoietic cells (CHs) and stromal multipotent mesenchymal (MSCs) are in close association studies aimed at the characterization of these may help to elucidate the mechanisms that govern tumor progression and identify novel therapeutic targets. Objective: To characterize and compare the CHs and MSCs derived from patients by conventional cytogenetics and molecular genes aurora kinase A and B. To evaluate the biological properties of MSCs derived from MDS and healthy controls. Methods: The study began with the clinical evaluation of 25 patients and eight healthy controls HCFMRP dooUSP and CH-Jau. Next, we performed a karyotypic analysis of CHs and MSCs from bone marrow by G-banding and FISH for aurora A and B genes and immunophenotypic profile and potential to differentiate into adipocytes and osteocytes of MSCs in patients with MDS and controls healthy. Results: The clinical evaluation showed thrombocytopenia (76%), neutropenia (100%), low hemoglobin (16%). The cytogenetic analysis revealed karyotype of CHs changed in 13 patients (52%), resulting in complex karyotype with numerical and structural changes. In contrast, in MSC, the karyotype was abnormal in seven patients (28%) and a pattern of lower complexity, only four patients had changes in both cell populations, however, different. Were found only numerical changes (monosomy being 86% and 14% gain in chromosome). The CHs and MSCs controls showed 100% normal karyotypes. In FISH analysis there was no evidence of gene amplification and AURKA AURKB. The MSCs of patients and controls were similar regarding the morphology and differentiation potential. However, the CTMs of patients proved to be changed to two surface antigens, CD90 and CD146, which showed higher expression levels in samples of patients (p = 0.04, p = 0.001 respectively). Conclusion: Furthermore, it was observed that the MSCs are changed although less frequently and differing from changes found in CHs. These data suggest that MSCs should play an important role in tumor progression and should be considered as targets in the search for new therapies and better explain the mechanisms that govern tumor progression. Although not shown AURKA amplification of genes in MDS and AURKB, future studies aimed at assessing the level of expression of these enzymes in patients with or without cytogenetic alterations may contribute to the understanding of the involvement or not of this gene with the disease. This study can not associate with profound anemia cytogenetic changes.

No description available.