O melanoma é uma lesão maligna da pele, com alta taxa de mortalidade cuja incidência vem aumentando nos últimos anos. Os principais fatores de risco são a história familial da doença, presença de nevos benignos múltiplos ou nevos atípicos e melanoma prévio. Imunossupressão, sensibilidade ao sol e exposição intermitente e intensa à radiação UV da luz solar, sem proteção, são fatores de risco adicionais. O objetivo deste estudo caso-controle de base hospitalar foi avaliar a contribuição de polimorfismos de genes de metabolização de xenobióticos (CYP1A1/MspI, CYP2E1/PstI, GSTM1, GSTT1 e GSTP1/Bsma), de genes de reparo do DNA (XRCC1/MspI, XRCC3/NcoI e XPD/PstI) e do gene do receptor de vitamina D (VDR/FokI e VDR/TaqI) no risco de melanoma. Consentiram em participar 193 pacientes com melanoma (49,7% homens e 50,3% mulheres, média de 52 ± 14,28 anos) e 208 controles (51,4% homens e 48,6% mulheres, média de 48 ± 15,24 anos) que após responderem a um questionário detalhado sobre hábitos e tipos de exposição a fatores de risco cederam amostras biológicas para análsie do DNA por PCR-RFLP. Os principais fatores de risco para o melanoma foram ascendência européia (p<0,001), cor de olhos claros (p<0,001), presença de nevos (p<0,001), histórico de queimadura grave na adolescência (p<0,001), falta de filtro solar (p<0.034) e exposição à lâmpadas fluorescentes (p=0,001). Quanto à análise dos polimorfismos de genes de metabolização de xenobióticos somente o GSTT1 nulo revelou associação inversamente positiva com o risco de melanoma maligno (OR ajustado = 0,60; IC95% = 0,37-0,97). Entretanto, essa associação não se manteve após a análise de regressão múltipla escalonada. Os polimorfismos VDR/FokI e VDR/TaqI não modificaram a susceptibilidade ao melanoma maligno na comparação entre os grupos. Na análise conjunta do fenótipo-genótipo, indivíduos com olhos verdes e genótipo VDR/FokI polimórfico, apresentaram risco praticamente seis vezes maior de melanoma (OR ajustado = 5,93; IC95% = 1,49-23,59). A associação entre os polimorfismos em pelo menos um dos alelos dos genes de reparo do DNA, XRCC3/NcoI e XPD/PstI aumentou praticamente duas vezes o risco de melanoma tanto na análise estatística multivariada (OR ajustado = 1,84; IC95% = 1,08-3,14) quanto na regressão logística múltipla escalonada (OR ajustado = 2,32; IC95% = 1,01-5,36). Na interação genemeio ambiente a falta do uso de filtro solar dobrou o risco de melanoma em indivíduos com polimorfismo XPD/PstI (OR ajustado = 2,17; IC95% = 1,12- 4,17). A identificação de polimorfismos genéticos associados com doenças multifatorias como o caso do melanoma maligno devem ser estimuladas em nosso meio, principalmente por vivermos em um país tropical com alta incidência solar em praticamente todo seu território. A identificação de marcadores genéticos de susceptibilidade pode propiciar medidas precoces e eficazes de prevenção do câncer. / Melanoma is a malignant skin lesion, with high mortalitty rate and its incidence has been rising in the last years. The main risk factors are melanoma family history, presence of multiple benign or atypical nevi and previous melanoma. Immunosuppression, sun sensitivy and intermittent and intense exposure to UV sunlight radiation, without protection, are additional risk factors. The aim of this hospital based case-control study was to evaluate the contribution of genetic polymorphisms of xenobiotic metabolizing enzymes (CYP1A1/MspI, CYP2E1/PstI, GSTM1, GSTT1 and GSTP1/Bsma), DNA repair genes (XRCC1/MspI, XRCC3/NcoI and XPD/PstI) and vitamin D receptor genes (VDR/FokI and VDR/TaqI) to the risk of melanoma. All participants, including 193 melanoma patients (49.7% men and 50.3% women, mean age 52 ± 14.28 years old) and 208 controls (51.4% men and 48.6% women, mean age 48 ± 15.24 years old) gave written informed consent to participate in the study and agreed to donate a sample of bloody to analysis of DNA by PCR-RFLP and answer a questionarie regarding phenotypic characteristics, personal habits and questions regarding sun exposure that could be associated to the disease. The main risk factors to melanoma were European ancestries (p<0.001), light colored eyes (p<0.001), presence of nevi (p<0.001), history of sunburns during the adolescence (p<0.001), no use of sunblock (p<0.034) and exposure of fluorescent lamps (p<0.001). Regarding the genes polymorphisms, only GSTT1 null genotype showed as an inversely positivefactor (OR adjusted = 0.60; 95%CI = 0.37-0.97) to malignant melanoma. However, this association disappeared with multiple regression analysis. The VDR/FokI and VDR/TaqI polymorphisms did not alter the susceptibility to malignant melanoma in the comparison between groups. A joint analysis of phenotype-genotype, individuals with green eyes and polymorphic genotype VDR/FokI, presented almost six times more risk to melanoma (OR adjusted = 5.93, 95% CI = 1.49-23.59). The association between polymorphisms, in at least one polymorphic allele of DNA repair genes XRCC3/NcoI and XPD/PstI increased almost twice the risk of melanoma in the multivariate statistic analysis (OR adjusted = 1.84, 95%CI = 1.08-3.14) and in multiple logistic regression (OR adjusted = 2.32, 95%CI = 1.01-5.36). In the interaction gene-environment the lack of sunscreen doubled the risk of melanoma in individuals with polymorphisms of XPD/PstI (OR adjusted = 2.17, 95%CI = 1.12-4.17). The identification of genetic polymorphisms associated with diseases such as multi factorial case of malignant melanoma should be encouraged in our country, mainly because we live in a tropical country with high solar irradiation in almost all its territory. The identification of genetic markers of susceptibility may provide early and effective prevention of cancer.

Submitted by Verena Bastos (verena@uefs.br) on 2015-09-21T13:18:11Z No. of bitstreams: 1 DISSERTACAO FINAL ALINE THOMAZ.pdf: 1680438 bytes, checksum: 85bf2d7886a394df745618b6fd50d435 (MD5) / Made available in DSpace on 2015-09-21T13:18:11Z (GMT). No. of bitstreams: 1 DISSERTACAO FINAL ALINE THOMAZ.pdf: 1680438 bytes, checksum: 85bf2d7886a394df745618b6fd50d435 (MD5) Previous issue date: 2015-09-27 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / The vitamin D receptor (VDR) is a cytoplasmic transcription factor and when activated by its ligand, translocates to the nucleus and interacts with DNA in the promoter regions with which has affinity, acting as a modulator of gene transcription and thereby producing multiple biological effects. Its main activities are the regulation and maintenance of plasma levels of calcium and phosphorus, as well as presenting immunomodulatory activities, such as the suppression of T cell activation, formation of secretion patterns of cytokines, modulation of proliferation and interference in the apoptosis. So this receptor is an important target for drugs that can help in the discovery of new immunomodulators. The present work had the objective to produce the recombinant vitamin D receptor in its soluble form for conducting future assays of drug screening of potential immunomodulators and drug-receptor interaction studies. For this, we used initially Escherichia coli expression system transformed with the plasmid HS_VDR_EC1-PQE T7, but it was only possible to obtain the protein in the insoluble fraction, even varying the temperature, time of induction and IPTG concentration. In an attempt to obtain soluble VDR, we used a eukaryotic expression system in insect cells using the baculovirus as a vector. It was built a vector, pFASTBacHT_VDR, which had the sequence of this protein cloned from pCMX.VDR. From there, it was possible to obtain recombinant bacmids used in transfection of insect cells, generating recombinant baculovirus, to then proceed with the expression of VDR. / O receptor de vitamina D (VDR) ? um fator de transcri??o g?nica citoplasm?tico e, quando ativado pelo seu ligante, transloca-se para o n?cleo e interage com regi?es promotoras no DNA com a qual apresenta afinidade, atuando como fator modulador da transcri??o g?nica e assim produzindo m?ltiplos efeitos biol?gicos. Suas principais atividades s?o a regula??o e a manuten??o dos n?veis plasm?ticos de c?lcio e f?sforo, e apresenta atividades imunomoduladoras, como a supress?o da ativa??o de c?lulas T, forma??o de padr?es de secre??o de citocinas, a modula??o da prolifera??o e interfer?ncia na apoptose. Sendo assim, esse receptor representa um importante alvo de drogas que pode contribuir na descoberta de novos f?rmacos com a??o imunomoduladora. O presente trabalho teve, como objetivo, produzir o VDR recombinante na forma sol?vel para a realiza??o de futuros ensaios de triagem de potenciais drogas com a??o imunomoduladora e estudos de intera??o droga-receptor. Para isso, foi utilizado, inicialmente, o sistema de express?o em Escherichia coli, utilizando o plasm?deo HS_VDR_EC1-PQE T7, por?m s? foi poss?vel obter a prote?na na fra??o insol?vel, mesmo variando a temperatura, o tempo de indu??o e a concentra??o de IPTG. Na tentativa de obter o VDR sol?vel, foi utilizado um sistema de express?o eucari?tico em c?lulas de inseto, utilizando como vetor o baculov?rus. Foi constru?do um vetor pFASTBacHT_VDR, o qual teve a sequ?ncia desta prote?na clonada a partir do pCMX.VDR. A partir da?, foi poss?vel obter bacm?deos recombinantes, utilizados na transfec??o de c?lulas de inseto, gerando baculov?rus recombinantes, para, ent?o, seguir com a express?o do VDR.

<p>In hyperparathyroidism (HPT) raised serum concentrations of ionized calcium is caused by increased secretion of parathyroid hormone (PTH) by parathyroid tumors. Active vitamin D, 1α,25-dihydroxyvitamin D₃, is known to suppress PTH secretion and to reduce proliferation of parathyroid tumor cells.</p><p>The aim of this thesis was to examine expression of vitamin D hydroxylating enzymes, regulating the activation and inactivation of vitamin D and to study effects of vitamin D analogues, in parathyroid tumors and breast cancer.</p><p>The vitamin D activating enzyme, CYP27B1/25-hydroxyvitamin D₃ 1α-hydroxylase (1α-hydroxylase) and the vitamin D inactivating enzyme CYP24A1/25-hydroxyvitamin D₃ 24-hydroxylase (24-hydroxylase) were expressed in parathyroid tumors and breast cancer. </p><p>The parathyroid tumors had raised expression levels of 1α-hydroxylase and reduced levels of 24-hydroxylase in comparison to normal parathyroid glands, indicating ability for endogenous activation of vitamin D. The expression of 1α-hydroxylase may be of therapeutic advantage for local activation of non-1α-hydroxylated vitamin D analogues in tumor cells, thereby reducing unwanted hypercalcemic effects. </p><p>Three of five selected low calcemic vitamin D analogues had as efficient PTH suppressing effect, in bovine parathyroid cells, as three vitamin D analogues used clinically for treatment of secondary HPT.</p><p>The non-1α-hydroxylated vitamin D analogue EB1285 showed antiproliferative and PTH suppressive effects as well as transcriptional activity in parathyroid and breast tumor cells, respectively.</p><p>Ketoconazole, an inhibitor of vitamin D hydroxylating enzymes, suppressed PTH secretion and potentiated the effect of vitamin D analogues. Combined treatment with vitamin D analogues and specific 24-hydroxylase inhibitors may be important for future therapy. </p>

<p>Important candidate genes to human bone phenotypes are those involved in the regulation of hormonal action, such as the vitamin D receptor (VDR) and the estrogen receptor-α (ERα) genes and their cofactors. RIZ1 is a specific ERα cofactor proved to strongly enhance the function of the ERα. </p><p>The main focus of this thesis has been to study genetic variants in the VDR and RIZ genes and their associations to human bone phenotypes using candidate gene and functional approaches. Specifically, polymorphisms in the VDR 3’ untranslated region (UTR) and a deletion/insertion polymorphism of a proline in the RIZ gene were investigated.</p><p>The candidate gene approach was applied to large-scale population-based cohorts of pre-and post-menopausal women from Sweden and of elderly men from Sweden and Hong Kong. VDR 3’ UTR polymorphisms were associated with peak bone mass and body composition in young women. Further analysis of common VDR 3’ UTR haplotypes confirmed the association with BMD and risk of fractures in elderly men from Sweden and Hong Kong. The VDR polymorphisms were investigated for cis-acting effects, affecting allelic expression in the normal chromosomal context of human bone cells. The VDR allelic transcripts in the bone samples were unequally expressed, suggesting presence of regulatory variants in the 3’ UTR. </p><p>The polymorphism in the RIZ gene was strongly associated to BMD in pre- and postmenopausal women and in elderly men. The functional analyses included reporter constructs containing the RIZ polymorphic variants transfected in a cell line and its abilities in coactivating the ERα were examined. The variants were functionally different in coactivating the ERα-receptor complex. </p><p>To summarize, the results of this thesis show novel evidence for functional relevant polymorphisms in candidate genes to human bone phenotypes. These polymorphisms may contribute to the variation seen in BMD and risk of fractures in the population.</p>

In hyperparathyroidism (HPT) raised serum concentrations of ionized calcium is caused by increased secretion of parathyroid hormone (PTH) by parathyroid tumors. Active vitamin D, 1α,25-dihydroxyvitamin D3, is known to suppress PTH secretion and to reduce proliferation of parathyroid tumor cells. The aim of this thesis was to examine expression of vitamin D hydroxylating enzymes, regulating the activation and inactivation of vitamin D and to study effects of vitamin D analogues, in parathyroid tumors and breast cancer. The vitamin D activating enzyme, CYP27B1/25-hydroxyvitamin D3 1α-hydroxylase (1α-hydroxylase) and the vitamin D inactivating enzyme CYP24A1/25-hydroxyvitamin D3 24-hydroxylase (24-hydroxylase) were expressed in parathyroid tumors and breast cancer. The parathyroid tumors had raised expression levels of 1α-hydroxylase and reduced levels of 24-hydroxylase in comparison to normal parathyroid glands, indicating ability for endogenous activation of vitamin D. The expression of 1α-hydroxylase may be of therapeutic advantage for local activation of non-1α-hydroxylated vitamin D analogues in tumor cells, thereby reducing unwanted hypercalcemic effects. Three of five selected low calcemic vitamin D analogues had as efficient PTH suppressing effect, in bovine parathyroid cells, as three vitamin D analogues used clinically for treatment of secondary HPT. The non-1α-hydroxylated vitamin D analogue EB1285 showed antiproliferative and PTH suppressive effects as well as transcriptional activity in parathyroid and breast tumor cells, respectively. Ketoconazole, an inhibitor of vitamin D hydroxylating enzymes, suppressed PTH secretion and potentiated the effect of vitamin D analogues. Combined treatment with vitamin D analogues and specific 24-hydroxylase inhibitors may be important for future therapy.

Important candidate genes to human bone phenotypes are those involved in the regulation of hormonal action, such as the vitamin D receptor (VDR) and the estrogen receptor-α (ERα) genes and their cofactors. RIZ1 is a specific ERα cofactor proved to strongly enhance the function of the ERα. The main focus of this thesis has been to study genetic variants in the VDR and RIZ genes and their associations to human bone phenotypes using candidate gene and functional approaches. Specifically, polymorphisms in the VDR 3’ untranslated region (UTR) and a deletion/insertion polymorphism of a proline in the RIZ gene were investigated. The candidate gene approach was applied to large-scale population-based cohorts of pre-and post-menopausal women from Sweden and of elderly men from Sweden and Hong Kong. VDR 3’ UTR polymorphisms were associated with peak bone mass and body composition in young women. Further analysis of common VDR 3’ UTR haplotypes confirmed the association with BMD and risk of fractures in elderly men from Sweden and Hong Kong. The VDR polymorphisms were investigated for cis-acting effects, affecting allelic expression in the normal chromosomal context of human bone cells. The VDR allelic transcripts in the bone samples were unequally expressed, suggesting presence of regulatory variants in the 3’ UTR. The polymorphism in the RIZ gene was strongly associated to BMD in pre- and postmenopausal women and in elderly men. The functional analyses included reporter constructs containing the RIZ polymorphic variants transfected in a cell line and its abilities in coactivating the ERα were examined. The variants were functionally different in coactivating the ERα-receptor complex. To summarize, the results of this thesis show novel evidence for functional relevant polymorphisms in candidate genes to human bone phenotypes. These polymorphisms may contribute to the variation seen in BMD and risk of fractures in the population.

Nuclear receptors (NRs) are ligand-activated transcription factors that regulate the expression of genes involved in all physiological activities. Disruption in NR function (e.g. mutations) can lead to a variety of diseases; making these receptors important targets for drug discovery. The ability to bind a broad range of 'drug-like' molecules also make these receptors attractive candidates for protein engineering, such that they can be engineered to bind novel small molecule ligands, for several applications. One application is the creation of potential molecular switches, tools that can be used for controlling gene expression. Gaining knowledge of specific molecular interactions that occur between a receptor and its ligand is of interest, as they contribute towards the activation or repression of target genes. The focus of this work has been to investigate the structural and functional relationships between the human vitamin D receptor (hVDR) and its ligands. To date, mutational assessments of the hVDR have focused on alanine scanning and residues typically lining the ligand binding pocket (LBP)that are involved in direct interactions with the ligand. A comprehensive analysis of the tolerance of these residues in the binding and activation of the receptor by its ligands has not been performed. Furthermore, residues not in contact with the ligand or that do not line the LBP may also play an important role in determining the activation profiles observed for NRs, and therefore need to be explored further. In order to engineer and use the hVDR in chemical complementation, a genetic selection system in which the survival of yeast is linked to the activation of a NR by an agonist, the hVDR gene was isolated from cDNA. To gain insight into how chemical and physical changes within the ligand binding domain (LBD) affect receptor-ligand interactions, libraries of hVDR variants exploring the role and tolerance of hVDR residues were created. To develop a comprehensive mutational analysis while also engineering the hVDR to bind a novel small molecule ligand, a rational and a random mutagenic approach were used to create the libraries. A variant, hVDRC410Y, that displayed enhanced activity with lithocholic acid (LCA), a known hVDR ligand, and novel activation with cholecalciferol (chole), a precursor of the hVDR's natural ligand known not to activate the wild-type hVDR, was discovered. The presence of a tyrosine at the C410 position resulting in novel activation profiles with both LCA and chole, and the fact that this residue does not line the hVDR's LBP led to interest in determining whether a physical or chemical property of the residue was responsible for the observed activity. When residue C410 was further assessed for its tolerance to varying amino acids, the results indicated that bulkiness at this end of the pocket is important for activation with these ligands. Both LCA and chole have reduced molecular volumes compared to the natural ligand, 1alpha, 25(OH)2D3. As a result, increased bulkiness at the C410 position may contribute additional molecular interactions between the receptor and ligands. Results obtained throughout this work suggest that the end of the hVDR's LBP consisting of two ligand anchoring residues, H305 and H397, and residue C410 tolerates structural variations, as numerous variants with mutations at these positions displayed enhanced activity. The receptor contains two tyrosines, Y143 and Y147, which were targeted for mutagenesis in one of the rationally designed libraries, located at the exact opposite end of the pocket. In an effort to gain further insight into the role of these residues at the other end of the LBP, mutagenesis assessing the tolerance of tyrosines 143 and 147 was performed. Overall, most changes at these positions proved to be detrimental to the function of the receptor supporting the hypothesis that this end of the LBP is less tolerant of structural changes, compared to the opposite end consisting of residues H305, H397 and C410. Overall, a better understanding of the structural and functional relationships between the human vitamin D receptor (hVDR) and its ligands was achieved. The effects of residue C410 on specificity and activation with the different ligands studied were unforeseen, as this residue does not line the receptor's ligand binding pocket (LBP). However, they serve as an example of the significant impact distant residues can have on receptor activation and also emphasize the important role physical properties of residues, such as volume, can play for specific ends of the LBP compared to chemical properties.

Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2016-10-18T17:32:34Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Tese Silvia Alves versão final biblioteca.pdf: 4384743 bytes, checksum: 6869052e5eb02362b089b2cba4aaff8b (MD5) / Made available in DSpace on 2016-10-18T17:32:34Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Tese Silvia Alves versão final biblioteca.pdf: 4384743 bytes, checksum: 6869052e5eb02362b089b2cba4aaff8b (MD5) Previous issue date: 2016-08-30 / CNPQ / A alergia às proteínas do leite de vaca (APLV) é definida como uma reação adversa imunologicamente mediada. A complexidade da APLV é observada tanto no espectro de apresentações clínicas como na sua história natural, e na sua etiologia, sobrepondo-se fatores genéticos e ambientais. O objetivo deste estudo foi analisar a frequência de polimorfismos nos genes das citocinas regulatórias (TGF-β1 e IL-10), inflamatória (IL-4) e no gene do receptor de vitamina D (VDR) em crianças com e sem APLV. A amostra foi selecionada por conveniência, com coleta de dados nos ambulatórios de Gastroenterologia Pediátrica, Alergia e Imunologia Pediátrica e Puericultura do Hospital das Clínicas da Universidade Federal de Pernambuco, entre abril de 2013 a junho de 2015. Foram avaliadas 154 crianças. Destas, 70 foram diagnosticadas com APLV (casos) e 84 eram saudáveis (grupo comparativo). O grupo com APLV apresentou maior proporção de crianças menores de um ano de idade (68,6% vs 35,7%; p=0,000), nascidos através de cesariana (85,7% vs 67,9%; p=0,010), com pais e mães com nível de escolaridade elevada e maior frequência de mães com atopia (55,7% vs 33,3%; p=0,005), em relação ao grupo comparativo. Dentre os casos, 31 (44,3%) foram classificados como portadores de sintomas predominantemente gastrintestinais e 39 (55,7%) com manifestação de sintomas cutâneos/respiratórios/anafilaxia. A idade (mediana) ao início dos sintomas foi menor no grupo gastrintestinal (30 dias vs 90 dias, p= 0,000), quando comparado ao grupo cutâneo/respiratório/anafilaxia. Ao término do estudo, a ausência de sintomas foi identificada em 29 (41,4%) das 70 crianças com APLV. A idade (mediana) nesse período foi menor no grupo gastrintestinal, em comparação ao grupo cutâneo/respiratório/anafilaxia (16 meses vs 20 meses; p= 0,036). A análise dos polimorfismos genéticos não apresentou diferença significante nas frequências genotípicas e alélicas para os genes das citocinas TGF-β1 (-509), IL-10 (-1082, -819 e -592), IL-4 (-590), receptor de IL-4 (IL-4R) e VDR (Fok I e Taq I) entre os casos e o grupo comparativo e entre os dois grupos de fenótipos clínicos e o grupo comparativo. Após controle para possíveis fatores de confusão, a cesariana (OR= 2,68; IC95%: 1,16 – 6,20) e a atopia materna (OR= 2,45; IC95%: 1,00 – 3,85) foram as variáveis associadas a maior chance para desenvolver APLV no grupo estudado. Quando estratificadas por fenótipo clínico, a menor duração do aleitamento materno exclusivo (OR= 4,85; IC95%: 1,81 – 13,03) e a atopia materna (OR= 3,59; IC95%: 1,44 – 8,92) foram associadas ao fenótipo clínico gastrintestinal, enquanto a cesariana (OR= 3,50; IC95%: 1,20 – 10,23) e o nascer prematuro (OR= 3,19; IC95%: 1,07 – 9,49) foram associados ao aumento da chance para o fenótipo clínico de reações cutâneas/respiratórias/anafilaxia. Concluiu-se que os polimorfismos nos genes das citocinas tolerogênicas (TGF-β1 e IL-10), inflamatória (IL-4) e do gene VDR (Fok I e Taq I) não apresentaram associação estatística com a APLV e os respectivos fenótipos clínicos. Contudo, a atopia materna, a menor duração do aleitamento materno exclusivo, a cesariana e a prematuridade apresentaram associação, podendo indicar que o ambiente pode exercer um efeito potencialmente maior sobre a patogênese dessa enfermidade. / Cow’s milk protein allergy (CMPA) is defined as an immunologically mediated adverse reaction. The complexity of CMPA is demonstrated both in the range of clinical presentations and its natural history as well as the etiology, with the overlapping of genetic and environmental aspects. The aim of the present study was to analyze the frequency of polymorphisms in the genes of regulatory (TGF-β1 and IL-10) and inflammatory (IL-4) cytokines as well as the gene of the vitamin D receptor (VDR) in children with and without CMPA. A convenience sample was used. Data were collected at the pediatric gastroenterology and pediatric allergy and immunology clinics of the hospital pertaining to the Federal University of Pernambuco (Brazil) between April 2013 and June 2015. A total of 154 children were evaluated, 70 of whom were diagnosed with CMPA (cases) and 84 were considered healthy (comparative group). The group with CMPA had higher proportions of children less than one year of age (68.6% vs. 35.7%; p=0.000), those born by cesarean section (85.7% vs. 67.9%; p=0.010), those whose parents had a high level of schooling and those whose mothers had atopy (55.7% vs. 33.3%; p=0.005) in comparison to the comparative group. Among the cases, 31 (44.3%) were classified as having predominantly gastrointestinal symptoms and 39 (55.7%) were diagnosed with skin/respiratory/anaphylaxis symptoms. Median age at the onset of symptoms was lower in the gastrointestinal group in comparison to the skin/respiratory/anaphylaxis group (30 days vs. 90 days, p=0.000). At the end of the study, an absence of symptoms was found in 29 of the 70 children with CMPA (41.4%). Median age in this period was lower in the gastrointestinal group than the skin/respiratory/anaphylaxis group (16 months vs. 20 months; p=0.036). The analysis of genetic polymorphisms revealed no significant differences in genotypic or allelic frequencies for the genes of the cytokines TGF-β1 (-509), IL-10 (-1082, -819 and -592), IL-4 (-590), IL-4 receptor (IL-4R) or vitamin D receptor-VDR (Fok I and Taq I) between the case and comparative groups or between the two clinical phenotype groups and comparative group. After controlling for possible confounding factors, cesarean delivery (OR=2.68; 95%CI: 1.16-6.20) and maternal atopy (OR=2.45; 95%CI: 1.00-3.85) were associated with a greater chance of developing CMPA. When stratified based on clinical phenotype, a shorter duration of exclusion breastfeeding (OR=4.85; 95%CI: 1.81-13.03) and maternal atopy (OR=3.59; 95%CI: 1.44-8.92) were associated with the gastrointestinal phenotype, whereas cesarean birth (OR=3.50; 95%CI: 1.20-10.23) and premature birth (OR=3.19; CI95%: 1.07-9.49) were associated with an increased chance of the clinical phenotype of skin/respiratory/anaphylaxis reactions. In conclusion, polymorphisms in the genes of tolerogenic (TGF-β1 and IL-10) and inflammatory (IL-4) cytokines and the VDR gene (Fok I and Taq I) were not significantly associated with CMPA or its clinical phenotypes. However, maternal atopy, a shorter duration of exclusion breastfeeding, cesarean delivery and premature birth were associated, which indicates that the environment can have a potentially greater effect on the pathogenesis of this disease.

Breast cancer is one of the leading causes of death in women cancer cases worldwide. Cancer is the result of environmental and genetic factors that contribute to alterations in cellular control, proliferation, differentiation and apoptosis. Vitamin D is emerging as an important nutrient in the prevention and treatment of cancer due to its ability to modulate proliferation and apoptosis in vivo and in vitro. To accomplish this, Vitamin D exerts its biological activity by binding to a specific, high-affinity intracellular vitamin D receptor (VDR). VDR expression is identified in mammary cancer cell lines, but levels are reduced compared to non-cancerous cells, which limits vitamin D-induced gene expression. Our study investigated two compounds with histone deacetylase inhibitor (HDACI) activity, trichostatin A (TSA), and sulforaphane (SFN), and how they influence the expression of vitamin D-induced gene expression. By isolating mRNA to create cDNA, we were able to run RT-PCR to analyze the overall gene expression. The genes investigated were: CYP24A1, CYP27B1, VDR and TRPV6. We found that in MCF-7 breast cancer cells, 1,25(OH)2D3 treatment alone induced the expression of VDR, CYP24A1 and CYP27B1. TRPV6 mRNA expression was not evident. TSA alone increased expression of VDR and CYP24A1, but SFN alone had no effect. Co-treatments of 1,25(OH)2D3 and TSA raised CYP24A1, but not significantly. Co-treatments with SFN seemed to decrease CYP24A1 expression, not significantly. Our findings support further study of the effects of the HDACI TSA in breast cancer, and suggest that this HDACI may be beneficial in augmenting vitamin D cellular responsiveness.

Breast cancer is a world health problem and is a leading cause of cancer-related death among women in the United States. However, breast cancer risks were reported to be reduced through exposure to Vitamin D through its Receptors identified as the p53 target gene. The purpose of this study was to assess the associations between VDR gene polymorphisms knowledge/awareness and decisions to reduce breast cancer risks and likelihood of mammogram screening among women in Texas. Data from survey were used. Roy adaptation model was the theoretical framework that guided this quasi- experimental, quantitative research. The dependent variables were decisions to reduce breast cancer risks and likelihood of mammogram screening. The independent variables were knowledge about VDR gene polymorphisms and exposure to vitamin D. The covariates were level of education, awareness, lifestyle, breast self-exams, mammograms, age, early menarche, late menopause, and family history of breast cancer. The chi-square test and regression analysis were used to test the stated research hypotheses and to answer the research questions. Knowledge of VDR gene polymorphisms and exposure to vitamin D were not significantly associated with breast cancer risk, ï?£2 (3, N= 250) =3.84, p > 0.05. Also, awareness of the risk factors for breast cancer was not significantly associated with decisions to go for mammogram screenings or to enroll in breast cancer risk-reduction programs, ï?£2 (3, N= 250) =1.58, p > 0.05. To advocate for the promotion of awareness of the importance of pharmacogenetic testing for VDR gene polymorphisms for early detection of breast cancer, which would help to undertake appropriate therapeutic measures in a timely manner to prevent cancer metastasis, further research is warranted.