A coenzima Q10 é uma provitamina lipossolúvel sintetizada endogenamente e naturalmente encontrada em alimentos como a carne vermelha, peixes, cereais, brócolis e espinafre. É comercializada como suplemento alimentar e utilizada em formulações cosméticas. Localiza-se na membrana de organelas celulares como retículo endoplasmático, vesículas e membrana interna da mitocôndria, onde atua como um cofator essencial na cadeia respiratória. Apresenta propriedades antioxidantes e potencial no tratamento de doenças neurodegenerativas e neuromusculares. O objetivo deste trabalho foi investigar os possíveis efeitos protetores de uma formulação hidrossolúvel de coenzima Q10 em células PC12 expostas à cisplatina, um fármaco antineoplásico que tem a neurotoxicidade como um dos fatores limitantes à sua utilização. A linhagem celular PC12 (feocromocitoma de ratos) utilizada nesta investigação é um reconhecido modelo in vitro para estudos neuronais. Os métodos empregados foram os ensaios do MTT, cometa, citoma micronúcleo com bloqueio da citocinese, crescimento de neuritos e análise da expressão do gene Tp53. Os resultados obtidos na avaliação da citotoxicidade da coenzima Q10 (0,1 - 20 µg/mL) mostraram que este antioxidante foi citotóxico às células PC12 na concentração de 20,0 µg/mL e não apresentou citotoxicidade em baixas concentrações. Para os ensaios do citoma e cometa, foram selecionadas três concentrações não citotóxicas de coenzima Q10 (0,1; 0,5 e 1,0 µg/mL) que não apresentaram mutagenicidade e genotoxicidade às células PC12. O efeito protetor da coenzima Q10 sobre a cisplatina no ensaio do citoma foi caracterizado pela diminuição da freqüência de micronúcleos e brotos nucleares, entretanto a proteção da coenzima Q10 não foi evidenciada no ensaio cometa. Alterações significativas na expressão do gene Tp53 não foram observadas no tratamento coenzima Q10 (1,0 µg/mL) associado à cisplatina (0,1 µg/mL). A coenzima Q10 (0,1 e 1,0 µg/mL) não foi neurotóxica em células PC12 indiferenciadas e diferenciadas após exposição ao fator de crescimento do nervo, e seu melhor efeito neuroprotetor foi observado na menor concentração avaliada. A coenzima Q10 reduziu a citotoxicidade da cisplatina (10,0 µg/mL) em células PC12 indiferenciadas e estimulou o crescimento de neuritos em células PC12 diferenciadas. A determinação dos efeitos citoprotetores da coenzima Q10 em um modelo neuronal é importante para elucidar possíveis estratégias de neuroproteção que poderiam ser aplicadas aos pacientes submetidos à quimioterapia. / Coenzyme Q10 is a liposoluble provitamin endogenously synthesized and naturally found in various foods items, such as meat, fish, cereals, broccoli and spinach. It is a dietary supplement in some countries and used in cosmetic formulations. Coenzyme Q10 is located in the membrane of cellular organelles such as endoplasmic reticulum, vesicles and inner mitochondrial membrane, where acts as an essential cofactor in the respiratory chain. It has antioxidant properties and potential in the treatment of neurodegenerative and neuromuscular diseases. The objective of this study was to investigate the possible protective effects of a water-soluble formulation of coenzyme Q10 in PC12 cells exposed to cisplatin, an anticancer drug that has neurotoxicity as a dose-limiting factor. The PC12 cell line (rat pheocromocytoma) used in this investigation is a recognized in vitro model for neuronal studies. The methods used were the MTT, comet, cytokinesis-block micronucleus cytome, neurite outgrowth assays and expression of Tp53 gene. The results obtained in the cytotoxicity of coenzyme Q10 (0.1-20 µg/mL) showed that this antioxidant was cytotoxic to PC12 cell at a concentration of 20.0 µg/mL and it was not cytotoxic at low concentrations. For the cytome and comet assays, were selected three non-cytotoxic concentrations of coenzyme Q10 (0.1, 0.5 and 1.0 µg/mL) without mutagenicity and genotoxicity PC12 cells. The protective effect of coenzyme Q10 in cytome assay was characterized by decreased frequency of micronuclei and nuclear buds induced by cisplatin, however the protection of coenzyme Q10 was not evidenced by the comet assay. No significant change in the Tp53 gene expression were observed in the coenzyme Q10 (1.0 µg/mL) plus cisplatin (0.1 µg/mL) treatment. Coenzyme Q10 (0.1 and 1.0 µg/mL) was not neurotoxic in undifferentiated and nerve growth factor differentiated PC12 cells and the lowest concentration evaluated showed the best neuroprotective effect. The coenzyme Q10 treatment reduced the citotoxicity of cisplatin (10.0 µg/mL) in undifferentiated PC12 cells and stimulated the neurite outgrowth in differentiated PC12 cells. Determination of the cytoprotective effects of the coenzyme Q10 in a neuronal model is important to elucidate possible strategies for neuroprotection that could be applied to patients undergoing chemotherapy.

Myelodysplastic syndrome (MDS) is a heterogeneous group of diseases characterized by ineffective hematopoiesis which is caused by damage of differentiation of pluripotent haematopoietic stem cells. TP53 gene mutations are identified approximately in 10% of MDS and represent a negative prognostic factor. Altered TP53 gene expression may have similar effect as the mutation. Mutations or deregulated expression of this gene have an impact on many cellular processes including apoptosis, DNA repair, cell growth and angiogenesis. In this work, the expression mRNA levels of genes involved in p53 signalling pathway were studied in CD34+ pluripotent haematopoietic cells from bone marrow of patients with low- risk MDS. MDS patients showed increased expression of genes involved in apoptosis induction, regulation of cell cycle and DNA repair (BAX, BBC3, CCNE1, CDC25A, CDKN1A, FAS, GADD45A) as compared to healthy subjects. The patients with TP53 mutation had decreased expression of apoptotic genes (BAX, PIDD, TRAF2) and increased gene expression of apoptotic inhibitor (BCL2A1), indicating a reduced activity of apoptotic pathways and that way the pathological cell clone may gain a growth advantage. Deregulation of 21 genes (BAX, BBC3, EGR1, KAT2B, MDM2 etc.) was observed in patients with del (5q) compared to...

Le carcinome hépatocellulaire (CHC) est la principale forme de cancer primitif du foie. Le CHC présente des variations d'incidence géographique qui reflètent les variations de prévalence des infections chroniques par les virus de l'hépatite B (VHB) et/ou C (VHC), ainsi que l'exposition alimentaire aux aflatoxines. Dans les régions de haute incidence, une mutation spécifique au le codon 249 du gène TP53 est très fréquemment détectée dans les CHC et a été proposée comme un marqueur moléculaire de l'exposition aux aflatoxines. La protéine mutée correspondante, p.R249S, interagit avec l'oncogène viral HBx, qui module la réplication virale, la prolifération et la survie cellulaire. Des séquences HBX sont détectables dans l'ADN génomique de plus de 80% des CHC liés au VHB. Nous avons étudié les associations de mutation R249S avec les caractéristiques moléculaires de HBX et la progression du CHC en utilisant des spécimens obtenus dans deux études cas-témoins développées Thaïlande et en Gambie. Nos résultats démontrent (1) que la mutation R249S est préférentiellement associée aux CHC qui se développent en l'absence de cirrhose hépatique préexistante ; (2) que la mutation est associée à la présence de polymorphismes dans l'intron 1 de TP53, suggérant l'influence de facteurs de susceptibilité génétique sur la formation des mutations ; (3) que la mutation est généralement associée à la rétention dans les cancers de séquences de HBX complètes. Ces résultats suggèrent que la protéine mutée p.R249S coopère avec HBx pour favoriser le développement de CHC sans cirrhose, et que la susceptibilité à la formation de cette mutation est influencée par la structure polymorphique de TP53 / Hepatocellular carcinoma (HCC) is the main form of primary liver cancer. HCC presents geographical variations in incidence that reflect variations in the prevalence of chronic infections by hepatitis B virus (HBV) and / or C (HCV) and dietary exposure to aflatoxins. In areas of high incidence, a specific mutation at codon 249 of TP53 gene is frequently detected in HCC and has been proposed as a molecular hallmark of aflatoxin exposure. The corresponding mutated protein p.R249S, interacts with the viral oncogene HBx, which modulates viral replication, proliferation and cell survival. HBX sequences are detectable in the genomic DNA of more than 80% of HBVrelated HCC. We investigated the associations of mutation R249S with the molecular characteristics of HBX and progression of HCC using specimens obtained in two case-control studies developed in Thailand and in The Gambia. Our results demonstrate (1) that the mutation R249S is preferentially associated with HCC that develop in the absence of pre-existing liver cirrhosis, (2) that the mutation is associated with the presence of polymorphisms in intron 1 of TP53, suggesting influence of genetic susceptibility factors on the formation of mutations, (3) that the mutation is usually associated with the retention of cancers with complete HBX sequences. These results suggest that the mutated protein p.R249S cooperates with HBx to promote the development of HCC without cirrhosis, and that susceptibility to the formation of this mutation is influenced by the polymorphic structure of TP53

Introdução: Embora o nascimento de gêmeos tenha sempre chamado atenção, não há fatores genéticos ou ambientais conhecidos que podem determinar o nascimento de monozigóticos gêmeos (MZ). E mesmo para gêmeos dizigóticos (DZ), influências genéticas não são completamente compreendidas. Objetivos: Um estudo prévio do nosso grupo demonstrou que o alelo C do polimorfismo rs1042522 no gene TP53 foi mais frequente nas mães de gêmeos do que nas mães de gestação única em uma pequena cidade no sul do Brasil. A fim de esclarecer se este foi um fator isolado, foi realizado um estudo observacional de caso-controle de base populacional. Métodos: As amostras foram selecionadas a partir de um programa de investigação de paternidade financiado pelo estado do Rio grande do Sul. As amostras foram consideradas casos em que duas das crianças tinham a mesma data de nascimento e os controles eram de amostras em que pelo menos dois filhos nasceram em datas diferentes. O método de escolha dos controles foi sequencial, sendo utilizados os primeiros que preenchessem esta condição a cada ano. Resultados: De 2007 a 2013, foram 32.661 registros pesquisados e 283 (0,9%) gêmeos foram encontrados (119 MZ e 164 DZ). Frequências alélicas e genotípicas não foram diferentes entre as mães de gêmeos ou mães de não gêmeos. No entanto, as mães de gêmeos MZ apresentaram uma maior frequência do genótipo GG e menor frequência do alelo C quando comparado com as mães de gêmeos DZ. Além disso, a proporção de gêmeos monozigóticos (42%) é maior do que normalmente relatado (30%). Finalmente, a proporção de gêmeos encontrados neste estudo parece ser mais realista, já que esta amostra não é supostamente de usuárias de técnicas de reprodução assistida. / Introduction: Although the birth of twins has always attracted attention, there are no known genetic or environmental factors that can determine the birth of monozygotic (MZ) twins. And even for dizygotic (DZ) twins, genetic influences are not completely understood. Objective: A previous study from our group has shown that the C allele of polymorphism rs1042522 in TP53 gene was more frequent in the mothers of twins than in the mothers of singletons in a small village in South Brazil. In order to clarify if this was an isolated factor, we performed a population-based observational case-control study. Methods: Samples were selected from a state-funded program of paternity investigation. Samples were considered cases when two of the children had the same date of birth whereas controls were those samples in which at least two children were born in different dates. The first subsequent sample fulfilling control criteria was included after each case. Results: From 2007 to 2013, 32,661 records were searched and 283 (0.9%) twins were found (119 MZ and 164 DZ). Genotypic and allele frequencies were not different between mothers of twins or mothers of singletons. However, mothers of MZ twins showed a higher frequency of GG genotype and lower frequency of the C allele when compared to mothers of DZ twins. Also, the proportion of MZ twins (42%) is higher than usually reported (30%). Finally, the proportion of twins found in this study seems to be more realistic, as this sample is allegedly not user of assisted reproduction techniques.

The tumor suppressor gene TP53 sits at the crux of response to cellular stresses. This is the most frequently inactivated gene in human tumors, being the target of somatic mutations. The protein product of TP53 is p53, and plays a crucial role in anti-proliferative signals through the induction of apoptosis, senescence, and cell-cycle arrest when activated by stresses such as genotoxic chemotherapeutic drugs. Therefore, the status of TP53 mutation in a tumor has profound implications for the tumorigenic potential as well as the response to anti-cancer therapies. Indeed, numerous studies have shown a predictive and prognostic value of TP53 mutations to the response to chemotherapy, but just as many studies show no significant contribution of TP53 mutations to chemotherapy response. This controversy is partly due to the lack of standard methods of TP53 mutation detection, but more importantly, it is due to the categorization of all TP53 mutations into one group. Certain mutations in TP53 can confer a mutant p53 with new, gained activities, not normally present in the WT p53 protein. These have been commonly called "gain of function" (GOF) p53 proteins, and some GOF p53 proteins can even confer oncogenic properties. However, not all gained functions are necessarily implicated in oncogenicity. Using stringent criteria, we have defined a select group of GOF TP53 mutations that do function as oncogenic proteins as oncomorphic TP53 mutations. In this work, we utilize data available from a large patient population through The Cancer Genome Atlas (TCGA) as well as data available from the University of Iowa Gynecologic Oncology Tumor Bank to examine the association of oncomorphic TP53 mutations with patient outcome using advanced serous ovarian cancer as a model. We demonstrate that oncomorphic TP53 mutations are associated with worse progression-free survival, chemoresistance, and higher rates of recurrence than other mutations in TP53 that have no evidence of oncomorphic abilities. We identify molecular alterations in patients with oncomorphic TP53 mutations, particularly the increased expression of β-catenin. We also observe that oncomorphic p53 proteins lose the normal protein:protein interactions with the microRNA microprocessing complex, implicating the role of dysregulated miRNAs in pathways associated with chemoresistance. The cumulative results from our studies provide human evidence for the consideration of different classes of TP53 mutations. Patients with oncomorphic TP53 mutations deserve careful follow-up therapy and may require novel treatment regimens to improve outcomes. We propose that stratification of patients should be considered based upon the individual TP53 mutation identified from their tumors.

La pèrdua de 17p13 succeeix en més del 70% dels tumors colorectals, i sòl acompanyar-se de mutació al locus TP53 de l'al·lel concomitant. A banda, hi ha prou indicis per a recolzar la hipòtesi que p53 és un marcador de pronòstic d'aquest tipus de tumors.A manca de consens sobre la millor estratègia per a conèixer el seu estat -no hi ha estàndard d'or-, ens proposàrem com a objectiu trobar un protocol aplicable a l'estudi rutinari de la proteïna p53 en el càncer colorectal. Per això, seleccionarem una sèrie prospectiva de casos d'adenocarcinoma primari, obtinguts per resecció quirúrgica, i d'ells recollirem, a banda dels resultats de la immunohistoquímica (IHQ) de p53, un seguit de dades clinicopatològiques: edat, sexe, mida del tumor, grau histològic, infiltració, afectació ganglionar i estadi clínic. Característiques demogràfiques de la sèrie foren: 100 tumors pertanyent a 96 pacients (38 dones i 58 homes), amb edat de presentació entre 37-93 anys. Els paràmetres obtinguts es resumeixen en: mida mitja dels tumors 4'1 cm, el 51% amb afectació ganglionar, grau histològic predominant moderadament diferenciat o grau 2 (81%) i nivell d'infiltració majoritari pT3 (72%).De l'abordatge IHQ en resultà que el 56% dels tumors evidencià acumulació de p53 (llindar de sensibilitat: 10%), mentre que un 29% dels casos negatius presentà manca total d'immunopositivitat. L'estudi del genotip, mitjançant PCR-SSCP/seqüenciació (exons 4-8), posà de manifest la presència de mutacions en el 68% dels tumors, el 73% de les quals conservaren la pauta de lectura. En analitzar les discrepàncies entre les 2 aproximacions concloguérem que el 39% dels casos fenotípicament negatius contenia mutacions al gen TP53, el que representa un 17% del total. Per contra, el 5% resultà mancat de mutacions -al menys, en els exons estudiats- malgrat presentar acumulació nuclear de p53. La representació mitjançant corba ROC dels resultats d'IHQ permeté afirmar que la tècnica s'ajusta a la de referència (SSCP/seqüenciació) en el càncer colorectal, amb una àrea sota la corba -equivalent de la precisió- del 79%. La regressió logística situà el llindar de sensibilitat òptim per a la IHQ en el 15%, percentatge en el qual es compensen millor els errors falsos, positius i negatius.La detecció de LOH del microsatèl·lit intragènic p53CA posà de manifest que el 63% dels tumors tenia pèrdua del locus TP53. L'anàlisi estadística demostrà una presència superior de mutacions en els tumors que havien perdut l'al·lel normal concomitant, i que els tumors d'estadi superior -en presència d'afectació ganglionar- tendien a presentar pèrdues de 17p.Conclusions: 1) la IHQ és un mètode vàlid per a cribrar alteracions de p53 en el càncer colorectal, malgrat requereixi ésser complementat amb estudis de SSCP/seqüenciació en els tumors de fenotip negatiu (llindar de sensibilitat: 15%); 2) el protocol d'aproximació a l'estat de p53 ha d'incloure la detecció de LOH al locus TP53. / More than 70% of colorectal adenocarcinomas exhibit 17p13 allelic losses, which usually occur with mutation at the TP53 locus of the concomitant allele. There are also studies that suggest that p53 is a prognostic marker for this type of tumors.Since there is no consensus about which is the best approach to assess p53 status -no gold standard has been defined, yet-, we undertook our study to establish a protocol suitable for the routine assessment of this marker in colorectal cancer. With this purpose, we selected a prospective series of primary adenocarcinomas, obtained by surgical resection and from them we compiled, apart from their p53 immunohistochemical results, the following clinicopathological data: age, sex, tumor size, histological grade, degree of invasion, nodal involvement and staging. Demographics of our series were: 100 tumors belonging to 96 patients (38 women and 58 men), 37-93 years of age at tumor presentation. Other parameters can be synthesized as: mean size of tumors 4.1 cm, 51% with nodal involvement, a prevalence of moderately differentiated tumors or grade 2 (81%), and the majority of tumors with pericolic fat invasion or pT3 (72%). The IHC approach allowed the detection of 56% of tumors with p53 protein accumulation (cutoff value: 10%), while up to 29% of the negative cases showed no immunopositivity at all. Genotype studies performed by PCR-SSCP/sequencing (exons 4-8) evidenced the presence of mutations in 68% of tumors, 73% of which preserved their gene reading frame. When discrepancies between the two approaches were analyzed, it was realized that 39% of the phenotypically negative cases were mutated at the TP53 gene, which represents 17% of total cases. On the contrary, 5% of tumors presenting nuclear accumulation of p53 protein resulted to be exempted of mutations, at least in the studied exons. The plot of IHC results by a ROC curve let us determine that this technique fitted to the reference one (SSCP/sequencing) for colorectal carcinoma, being the area under the curve -equivalent to accuracy- of 79%. Logistic regression established the optimal cutoff value at 15%, percentage at which false positive and negative errors are best balanced.Detection of LOH using the intragenic p53CA dinucleotide microsatellite revealed that 63% of tumors had allelic loss at the TP53 locus. Statistical analyses demonstrated a superior presence of mutations in tumors that had lost its wildtype concomitant allele, and also that tumors with higher staging, based on nodal involvement, tended to present 17p allelic loss.Conclusions: 1) IHC is a valid method to screen p53 alterations in colorectal cancer, although it is necessary that negative tumors (cutoff value: 15%) are also complemented by SSCP/sequencing. 2) A protocol to assess p53 status must include the detection of LOH at the TP53 locus.

The goniothalamin (GTN) and related styryl-lactones were found to be cytotoxic and apoptotic to a variety of tumor cell lines including breast cancer MCF-7, cervical cancer HeLa and leukemia HL60. In HL60 cells, GTN triggers mitochondria dysfunction, caspase activation and eventually, apoptosis. In this study, we demonstrated that GTN was able to induce apoptosis in hepatocellular carcinoma derived cells, SK-Hep1 and Hep-3B cells via upregulation of the phorbol-12-myristate-13-induced 1 (PMAIP1) protein , alternation of mitochondrial membrane potentials (P < 0.05) via TP53-dependent and -independent transactivations. Treatment with GTN induced DNA damage, formation of reactive oxygen species (ROS, P < 0.05) and activated cleaved CASP8, CASP9, CASP3 and poly (ADP-ribose) polymerase 1 proteins. However, cleaved BH3 interacting domain (BID) death agonist protein was not identified, suggesting that an intrinsic cellular stress was produced after GTN treatments in both cell lines. A pan caspase inhibitor, z-VAD-FMK, suppressed GTN-induced apoptotic cell percentages (P < 0.05), demonstrating that GTN-induced apoptosis was mediated by the activation of the caspase cascade protein. The GTN induced ROS formation prior to DNA damage in SK-Hep1, yet in reverse order in Hep-3B cells. Moreover, GTN induced DNA damages through activation of the gamma H2A histone family member X (£^H2AFX, Ser139)/phospho-CHK1 checkpoint homolog (pCHEK1, Ser345)/phospho-CHK2 checkpoint homolog (pCHEK2, Thr68)/phosphos-tumor protein p53 (pTP53, Ser15), and £^H2AFX/pCHEK2 (Thr68), in SK-Hep1 and Hep-3B cells,respectively. Among five integral outer mitochondrial membrane proteins that blocks or induces apoptotic death, PMAIP1 protein and PMAIP1 mRNA levels were upregulated after GTN treatments for 4 to 6 h in both cell lines (P < 0.05). Transfection of shRNA interference plasmids targeting PMAIP1 gene downregulated PMAIP1 mRNA (P < 0.05) and PMAIP1 protein (P < 0.05) levels, as well as GTN-induced apoptotic cell percentages (P < 0.05) in both cell lines. In parallel, transfection of the shRNA interference plasmid targeting TP53 gene in SK-Hep1 cells, downregulated TP53 and PMAIP1 mRNA (P < 0.05) and TP53, pTP53, PMAIP1, cleaved PARP1 protein levels and apoptotic cell percentages (P < 0.05). Treatment with the TP53 inhibitor, PFT-£\ or transfection of a dominant negative TP53 plasmid, pTP53mt135, repressed TP53, pTP53 and PMAIP1 protein and/or TP53, PMAIP1 mRNA levels (P < 0.05), however, significantly augmented GTN-induced apoptotic cell percentages (P < 0.05). Cytosol/mitochondria fractionation identified that TP53, along with PMAIP1 proteins, were translocated to mitochondria after GTN treatment in time-dependent manners. Taken together, our studies demonstrated that GTN induced apoptosis via PMAIP1 via both TP53-dependent and -independent transactivation pathways. In TP53-positive SK-Hep1 cells, PMAIP1 and TP53 proteins conducted synergic effects.

The objective was to study apoptotic effects and underlying molecular mechanisms of goniothalamin (GTN) in hepatocellular carcinoma (HCC)-derived cells. The GTN that isolated from Goniothalamus amuyon, was found to possess profound cytotoxic activities against human SK-Hep1 and Hep-3B cells in vitro. The cytotoxicity of GTN/cell viability was measured by MTT assay at 570-nm absorbance and the IC50 at 72 h were determined as 7.5, 17 mM in SK-Hep1 and Hep-3B respectively. The GTN induced cell death and accumulation of reactive oxygen species in HCC-derived cells. One reactive oxygen species inhibitor, N-actylcysteine, further restored cell viabilities post-GTN treatments. Formation of £^-H2AX foci suggested that GTN-induced DNA damages were double-strand breaks. The GTN arrested cell cycle at G0/G1 by regulation of CCND, CCNE1, RB1 and E2F1 proteins in SK-Hep1, and at G2/M by regulation of CCNB1 in Hep-3B cells. The GTN-induced apoptosis in HCC-derived cells were evidenced by phosphatidylserine externalization and involved both extrinsic and intrinsic pathways in SK-Hep1 cells, but only extrinsic pathway in Hep-3B cells. In SK-Hep1 cells, GTN induced apoptosis and cell cycle arrest through TP53-mediated pathway in contrast to that of TP53/FAS mutated Hep-3B cells. Importantly, GTN was able to induce apoptosis in both TP53 wild type and TP53/FAS mutated HCC-derived cells. On the other hand, GTN was able to induce TP53 and p21WAF1/Cip1 up-regulation in SK-Hep1 and Hep-3B cells and p27Kip1 up-regulation in Hep-3B cells. These results demonstrated that GTN induced apoptosis in HCC cells through distinct signaling pathways.

Introdução: Embora o nascimento de gêmeos tenha sempre chamado atenção, não há fatores genéticos ou ambientais conhecidos que podem determinar o nascimento de monozigóticos gêmeos (MZ). E mesmo para gêmeos dizigóticos (DZ), influências genéticas não são completamente compreendidas. Objetivos: Um estudo prévio do nosso grupo demonstrou que o alelo C do polimorfismo rs1042522 no gene TP53 foi mais frequente nas mães de gêmeos do que nas mães de gestação única em uma pequena cidade no sul do Brasil. A fim de esclarecer se este foi um fator isolado, foi realizado um estudo observacional de caso-controle de base populacional. Métodos: As amostras foram selecionadas a partir de um programa de investigação de paternidade financiado pelo estado do Rio grande do Sul. As amostras foram consideradas casos em que duas das crianças tinham a mesma data de nascimento e os controles eram de amostras em que pelo menos dois filhos nasceram em datas diferentes. O método de escolha dos controles foi sequencial, sendo utilizados os primeiros que preenchessem esta condição a cada ano. Resultados: De 2007 a 2013, foram 32.661 registros pesquisados e 283 (0,9%) gêmeos foram encontrados (119 MZ e 164 DZ). Frequências alélicas e genotípicas não foram diferentes entre as mães de gêmeos ou mães de não gêmeos. No entanto, as mães de gêmeos MZ apresentaram uma maior frequência do genótipo GG e menor frequência do alelo C quando comparado com as mães de gêmeos DZ. Além disso, a proporção de gêmeos monozigóticos (42%) é maior do que normalmente relatado (30%). Finalmente, a proporção de gêmeos encontrados neste estudo parece ser mais realista, já que esta amostra não é supostamente de usuárias de técnicas de reprodução assistida. / Introduction: Although the birth of twins has always attracted attention, there are no known genetic or environmental factors that can determine the birth of monozygotic (MZ) twins. And even for dizygotic (DZ) twins, genetic influences are not completely understood. Objective: A previous study from our group has shown that the C allele of polymorphism rs1042522 in TP53 gene was more frequent in the mothers of twins than in the mothers of singletons in a small village in South Brazil. In order to clarify if this was an isolated factor, we performed a population-based observational case-control study. Methods: Samples were selected from a state-funded program of paternity investigation. Samples were considered cases when two of the children had the same date of birth whereas controls were those samples in which at least two children were born in different dates. The first subsequent sample fulfilling control criteria was included after each case. Results: From 2007 to 2013, 32,661 records were searched and 283 (0.9%) twins were found (119 MZ and 164 DZ). Genotypic and allele frequencies were not different between mothers of twins or mothers of singletons. However, mothers of MZ twins showed a higher frequency of GG genotype and lower frequency of the C allele when compared to mothers of DZ twins. Also, the proportion of MZ twins (42%) is higher than usually reported (30%). Finally, the proportion of twins found in this study seems to be more realistic, as this sample is allegedly not user of assisted reproduction techniques.

Mutações germinativas no gene TP53 estão associadas com a Síndrome de Li-Fraumeni e sua variante, a Síndrome de Li-Fraumeni-Like, ambas doenças autossômicas dominantes caracterizadas pela predisposição a múltiplos tumores em idade precoce. Recentemente p53 foi descrita como uma proteína com funções antioxidantes, atuando na manutenção da estabilidade genômica. Sendo assim, o objetivo deste trabalho foi investigar parâmetros de estresse oxidativo em sangue periférico de indivíduos portadores da mutação germinativa TP53 p.R337H em comparação com indivíduos não portadores da alteração. Foi verificado que a atividade eritrocitária da enzima GPx, uma importante defesa antioxidante, diferiu significativamente entre portadores e não portadores da mutação, com um aumento da atividade nos primeiros (p=0.048). O conteúdo de grupamentos carbonila e de malondialdeído, indicadores de dano à proteína e a lipídios, respectivamente, também foram significativamente maiores no grupo dos portadores (p=0.04 e p<0.0001). Por fim, indivíduos portadores da mutação apresentaram um aumento no estado antioxidante total, mas uma diminuição no conteúdo de ácido ascórbico em plasma. Nossos resultados apontam para alterações da função antioxidante de p53 em indivíduos portadores da mutação germinativa TP53 p.R337H, independentemente da idade e do diagnóstico prévio de um tumor. Estas alterações podem estar diretamente relacionadas à predisposição ao câncer neste grupo de pacientes. / Germline mutations in the TP53 gene are the underlying genetic defect of Li-Fraumeni Syndrome (LFS) and its variant, Li-Fraumeni-like (LFL) Syndrome, autosomal dominant disorders characterized by predisposition to multiple early-onset cancers. More recently, p53 is emerging as an important player in redox metabolism and important enzymes involved in defenses against oxidative stress have shown to be regulated by p53, including glutathione peroxidase 1 (GPX1) and mitochondrial superoxide dismutase 2 (SOD2). The aim of the present study was to investigate the redox profile parameters in blood of p.R337H mutation carriers and non-carriers individuals. A total of 34 individuals were included in the study and they were divided in two groups: mutation carriers (C) (n=17) and non-carriers (NC) (n=17). Erythrocyte GPx activity, an important enzymatic antioxidant defense, differed significantly between carriers and non carriers, with an increased activity in the latter (p = 0.048). Plasma Carbonyl content, an indicative of protein damage related to ROS overgeneration, was also increased in carriers (p = 0.04). Furthermore, an association between malondialdehyde (MDA, a lipidic peroxidation product) levels and mutation status was found in plasma. Mutation carriers had a four-fold increase in MDA levels (C= 40.20 ± 2.84, NC=160.5 ± 3.64, p < 0.0001) when compared to non carriers. Finally, mutation carriers showed an increased total antioxidant status (TAS), but a decrease in the Vitamin C content in plasma, when compared to non-carriers. Our findings suggest that TP53 p.R337H mutation carriers present an imbalance in oxidative metabolism parameters, which could be directly related to the pathophysiological mechanisms of cancer predisposition in these individuals.