Epigenetic Disruption of the Piwi Pathway in Human Spermatogenic Disorders

Epigenetic changes are involved in a wide range of common human diseases. Although DNA methylation defects are known to be associated with male infertility in mice, their impact on human deficiency of sperm production has yet to be determined. We have assessed the global genomic DNA methylation profiles in human infertile male patients with spermatogenic disorders by using the Infinium Human Methylation27 BeadChip. Three populations were studied: conserved spermatogenesis, spermatogenic failure due to germ cell maturation defects, and Sertoli cell-only syndrome samples. A disease-associated DNA methylation profile, characterized by targeting members of the PIWI-associated RNA (piRNA) processing machinery, was obtained. Bisulfite genomic sequencing and pyrosequencing in a large cohort (n = 46) of samples validated the altered DNA methylation patterns observed in piRNA-processing genes. In particular, male infertility was associated with the promoter hypermethylation-associated silencing of PIWIL2 and TDRD1. The downstream effects mediated by the epigenetic inactivation of the PIWI pathway genes were a defective production of piRNAs and a hypomethylation of the LINE-1 repetitive sequence in the affected patients. Overall, our data suggest that DNA methylation, at least that affecting PIWIL2/TDRD1, has a role in the control of gene expression in spermatogenesis and its imbalance contributes to an unsuccessful germ cell development that might explain a group of male infertility disorders

BMP Inhibition in Seminomas Initiates Acquisition of Pluripotency via NODAL Signaling Resulting in Reprogramming to an Embryonal Carcinoma

Type II germ cell cancers (GCC) can be subdivided into seminomas and non-seminomas. Seminomas are similar to carcinoma in situ (CIS) cells, the common precursor of type II GCCs, with regard to epigenetics and expression, while embryonal carcinomas (EC) are totipotent and differentiate into teratomas, yolk-sac tumors and choriocarcinomas. GCCs can present as seminomas with a non-seminoma component, raising the question if a CIS gives rise to seminomas and ECs at the same time or whether seminomas can be repro- grammed to ECs. In this study, we utilized the seminoma cell line TCam-2 that acquires an EC-like status after xenografting into the murine flank as a model for a seminoma to EC tran- sition and screened for factors initiating and driving this process. Analysis of expression and DNA methylation dynamics during transition of TCam-2 revealed that many pluripotency- and reprogramming-associated genes were upregulated while seminoma-markers were downregulated. Changes in expression level of 53 genes inversely correlated to changes in DNA methylation. Interestingly, after xenotransplantation 6 genes ( GDF3 , NODAL , DNMT3B , DPPA3 , GAL , AK3L1 ) were rapidly induced, followed by demethylation of their genomic loci, suggesting that these 6 genes are poised for expression driving the repro- gramming. We demonstrate that inhibition of BMP signaling is the initial event in reprogram- ming, resulting in activation of the pluripotency-associated genes and NODAL signaling. We propose that reprogramming of seminomas to ECs is a multi-step process. Initially, the microenvironment causes inhibition of BMP signaling, leading to induction of NODAL sig- naling. During a maturation phase, a fast acting NODAL loop stimulates its own activity and temporarily inhibits BMP signaling. During the stabilization phase, a slow acting NODAL loop, involving WNTs re-establishes BMP signaling and the pluripotency circuitry. In parallel, DNMT3B-driven de novo methylation silences seminoma-associated genes and epigenetically fixes the EC state

DNMT3A mutations mediate the epigenetic reactivation of the leukemogenic factor MEIS1 in acute myeloid leukemia

Close to half of de novo acute myeloid leukemia (AML) cases do not exhibit any cytogenetic aberrations. In this regard, distortion of the DNA methylation setting and the presence of mutations in epigenetic modifier genes can also be molecular drivers of the disease. In recent years, somatic missense mutations of the DNA methyltransferase 3A (DNMT3A) have been reported in ~20% of AML patients; however, no obvious critical downstream gene has been identified that could explain the role of DNMT3A in the natural history of AML. Herein, using whole-genome bisulfite sequencing and DNA methylation microarrays, we have identified a key gene undergoing promoter hypomethylation-associated transcriptional reactivation in DNMT3 mutant patients, the leukemogenic HOX cofactor MEIS1. Our results indicate that, in the absence of mixed lineage leukemia fusions, an alternative pathway for engaging an oncogenic MEIS1-dependent transcriptional program can be mediated by DNMT3A mutations

Circular RNA CpG island hypermethylation-associated silencing in human cancer

Noncoding RNAs (ncRNAs), such as microRNAs and long noncoding RNAs (lncRNAs), participate in cellular transformation. Work done in the last decade has also demonstrated that ncRNAs with growth-inhibitory functions can undergo promoter CpG island hypermethylation-associated silencing in tumorigenesis. Herein, we wondered whether circular RNAs (circRNAs), a type of RNA transcripts lacking 5′-3′ ends and forming closed loops that are gaining relevance in cancer biology, are also a target of epigenetic inactivation in tumors. To tackle this issue, we have used cancer cells genetically deficient for the DNA methyltransferase enzymes in conjuction with circRNA expression microarrays. We have found that the loss of DNA methylation provokes a release of circRNA silencing. In particular, we have identified that promoter CpG island hypermethylation of the genes TUSC3 (tumor suppressor candidate 3), POMT1 (protein O-mannosyltransferase 1), ATRNL1 (attractin-like 1) and SAMD4A (sterile alpha motif domain containing 4A) is linked to the transcriptional downregulation of both linear mRNA and the hosted circRNA. Although some circRNAs regulate the linear transcript, we did not observe changes in TUSC3 mRNA levels upon TUSC3 circ104557 overexpression. Interestingly, we found circRNA-mediated regulation of target miRNAs and an in vivo growth inhibitory effect upon TUSC3 circ104557 transduction. Data mining for 5′-end CpG island methylation of TUSC3, ATRNL1, POMT1 and SAMD4A in cancer cell lines and primary tumors showed that the epigenetic defect was commonly observed among different tumor types in association with the diminished expression of the corresponding transcript. Our findings support a role for circRNA DNA methylation-associated loss in human cancer

VASCULAR AND STRUCTURAL PULP CHANGES INDUCED BY ORTHODONTIC FORCES: A REVIEW

It is thought that, in occasions, the orthodontic forces produce alterations in the angiogenesis, structures and capillary pulpar flow. Nevertheless, it can be found reports of stimulation of the angiogenesis through the growth factors; structural changes due to fibrosis, apoptosis, pulpar degenerations and necrosis; vascular changes by a reduction in the sanguineous flow to pulpar, and changes in the trustworthiness of the sensitivity tests. The purpose of this article is to raise several questions on the structural and vascular pulpar changes induced by orthodontic forces.Se ha sugerido que, en ocasiones, las fuerzas ortodónticas producen alteraciones en la angiogénesis, estructura y flujo sanguíneo pulpar. Sin embargo, en la literatura se encuentran reportes de estimulación de la angiogénesis a través de los factores de crecimiento; cambios estructurales debidos a fibrosis, apoptosis, degeneraciones pulpares y necrosis; cambios vasculares por una reducción en el flujo sanguíneo pulpar, y cambios en la confiabilidad de las pruebas de sensibilidad. El propósito de esta revisión es plantear varios cuestionamientos sobre los cambios pulpares vasculares y estructurales inducidos por fuerzas ortodónticas.[Delgado L, Ojeda CA, Ferreira H, Ordoñez E. Cambios pulpares vasculares y estructurales inducidos por fuerzas ortodónticas: Una revisión. Ustasalud Odontología 2005: 4: 44 – 47

Study on antibacterial activity of silver nanoparticles synthesized by gamma irradiation method using different stabilizers

In order to observe the microbiological status of CMT positive samples, 734 apparently health mammary quarters from buffalo cows were submitted to physical evaluation, strip cup test and CMT. After milk samples inoculation in 10% ovine blood agar base media and in MacConkey agar and incubation under aerobic condition for 72 hours at 37 degrees C, identification was proceeded. According to CMT, 227 quarters (30,93%) were positive, among them 73 (32,16%) presented 1+ reaction, 53 (23,35%) were 2+ and 101 (44,49%) were 3+. Microbiological exams of such samples were positive in 147 (64,76%) out of 227 CMT positive samples and among the remaining 72 (31,72%) were negative and 8 (3,52) were contaminated. In the 147 microbiological positive samples 204 bacteria were found in pure or associated growth and the most frequent agents were: Corynebacterium sp (59,25%); Staphylococcus sp (17,65%) among which 86,11% were coagulase negative and 13,89% were coagulase positive; and Micrococcus sp (6,37%). The results revealed that, excluding the eight contaminated samples, 147 (67,12%) quarters out of 219 CMT positive could be considered as bacteria-carrier and that even in a smaller percentage false-positive results can cause problems in a sanitary program for mastitis control in dairy buffalo cows

Epigenetic silencing of TGFBI confers resistance to trastuzumab in human breast cancer

Background: acquired resistance to trastuzumab is a major clinical problem in the treatment of HER2-positive (HER2+) breast cancer patients. The selection of trastuzumab-resistant patients is a great challenge of precision oncology. The aim of this study was to identify novel epigenetic biomarkers associated to trastuzumab resistance in HER2+ BC patients. Methods: we performed a genome-wide DNA methylation (450K array) and a transcriptomic analysis (RNA-Seq) comparing trastuzumab-sensitive (SK) and trastuzumab-resistant (SKTR) HER2+ human breast cancer cell models. The methylation and expression levels of candidate genes were validated by bisulfite pyrosequencing and qRT-PCR, respectively. Functional assays were conducted in the SK and SKTR models by gene silencing and overexpression. Methylation analysis in 24 HER2+ human BC samples with complete response or non-response to trastuzumab-based treatment was conducted by bisulfite pyrosequencing. Results: epigenomic and transcriptomic analysis revealed the consistent hypermethylation and downregulation of TGFBI, CXCL2, and SLC38A1 genes in association with trastuzumab resistance. The DNA methylation and expression levels of these genes were validated in both sensitive and resistant models analyzed. Of the genes, TGFBI presented the highest hypermethylation-associated silencing both at the transcriptional and protein level. Ectopic expression of TGFBI in the SKTR model suggest an increased sensitivity to trastuzumab treatment. In primary tumors, TGFBI hypermethylation was significantly associated with trastuzumab resistance in HER2+ breast cancer patients. Conclusions: our results suggest for the first time an association between the epigenetic silencing of TGFBI by DNA methylation and trastuzumab resistance in HER2+ cell models. These results provide the basis for further clinical studies to validate the hypermethylation of TGFBI promoter as a biomarker of trastuzumab resistance in HER2+ breast cancer patients

An insight into the sialotranscriptome and virome of Amazonian anophelines

Background: Saliva of mosquitoes contains anti-platelet, anti-clotting, vasodilatory, anti-complement and anti-inflammatory substances that help the blood feeding process. The salivary polypeptides are at a fast pace of evolution possibly due to their relative lack of structural constraint and possibly also by positive selection on their genes leading to evasion of host immune pressure. Results: In this study, we used deep mRNA sequence to uncover for the first time the sialomes of four Amazonian anophelines species (Anopheles braziliensis, A. marajorara, A. nuneztovari and A. triannulatus) and extend the knowledge of the A. darlingi sialome. Two libraries were generated from A. darlingi mosquitoes, sampled from two localities separated ~ 1100 km apart. A total of 60,016 sequences were submitted to GenBank, which will help discovery of novel pharmacologically active polypeptides and the design of specific immunological markers of mosquito exposure. Additionally, in these analyses we identified and characterized novel phasmaviruses and anpheviruses associated to the sialomes of A. triannulatus, A. marajorara and A. darlingi species. Conclusions: Besides their pharmacological properties, which may be exploited for the development of new drugs (e.g. anti-thrombotics), salivary proteins of blood feeding arthropods may be turned into tools to prevent and/or better control vector borne diseases; for example, through the development of vaccines or biomarkers to evaluate human exposure to vector bites. The sialotranscriptome study reported here provided novel data on four New World anopheline species and allowed to extend our knowledge on the salivary repertoire of A. darlingi. Additionally, we discovered novel viruses following analysis of the transcriptomes, a procedure that should become standard within future RNAseq studies. © 2019 The Author(s)

Relação do índice de desenvolvimento humano e as variáveis nutricionais em crianças do brasil

Objetivo O objetivo do presente estudo é demonstrar a relação do Indice de desenvolvimento humano (IDH) de diferentes regiões brasileiras no comportamento das variáveis de composicão corporal-indicadores do estado nutricional, assim como, a adequação da utilização destas variáveis.Materiais e Método Estudo descritivo, de corte transversal de topología comparativa. A amostra foi composta por alunos de escolas públicas, de ambos os sexos, com idade entre 8 e 10 anos, de 3 regiões brasileiras, escolhidas de maneira aleatória, sendo: Sul n= 262 masculino e n=251 feminino; Nordeste n=45 masculino y n=35 feminino; Norte n=96 masculino y n=38 feminino.Os protocolos utilizados foram de estatura, massa, somatória de dobras cutáneas e IMC. O IDH foi retirado do Programa das Nações Unidas para o Desenvolvimento. A estatística utilizada foi a descritiva e inferencial, através do método comparativo Anova one-way para os dados paramétricos e o teste Kurskal-Wallis para os dados não paramétricos, sendo adotado um nível de significância de p and lt;0,05, ou seja, 95 % de probabilidade para as afirmativas e/ou negativas, denotadas durante as investigações.ResultadosFoi demostrada a existência de uma diferencia significativa p and lt;0.0001 nas variáveis estudadas.Conclusiones A utilização de antropometria como uma ferramenta auxiliar para conhecer a composição corporal das crianças podem ajudar na prevenção ou tratamento precoce dos distúrbios alimentares, como desnutrição e obesidade. Ela pode ser parte das intervenções das políticas públicas ligadas à assistência infantil,na utilização de formas simples e eficiente para evitar problemas de saúde pública

Distinct DNA methylomes of newborns and centenarians

Human aging cannot be fully understood in terms of the constrained genetic setting. Epigenetic drift is an alternative means of explaining age-associated alterations. To address this issue, we performed whole-genome bisulfite sequencing (WGBS) of newborn and centenarian genomes. The centenarian DNA had a lower DNA methylation content and a reduced correlation in the methylation status of neighboring cytosine--phosphate--guanine (CpGs) throughout the genome in comparison with the more homogeneously methylated newborn DNA. The more hypomethylated CpGs observed in the centenarian DNA compared with the neonate covered all genomic compartments, such as promoters, exonic, intronic, and intergenic regions. For regulatory regions, the most hypomethylated sequences in the centenarian DNA were present mainly at CpG-poor promoters and in tissue-specific genes, whereas a greater level of DNA methylation was observed in CpG island promoters. We extended the study to a larger cohort of newborn and nonagenarian samples using a 450,000 CpG-site DNA methylation microarray that reinforced the observation of more hypomethylated DNA sequences in the advanced age group. WGBS and 450,000 analyses of middle-age individuals demonstrated DNA methylomes in the crossroad between the newborn and the nonagenarian/centenarian groups. Our study constitutes a unique DNA methylation analysis of the extreme points of human life at a single-nucleotide resolution level