LINE-1 methylation in cleft lip tissues:influence of infant MTHFR c.677C>T genotype

Objective: To investigate the influence of MTHFR c.677C>T genotype on LINE-1 methylation in lateral and medial tissues from cleft lip (CL). Methods: Forty-five consecutive non-syndromic cleft lip with or without cleft palate (nsCL/P) cases were included in the study. Genomic DNA was extracted from tissues at both sides of cleft lip, and LINE-1 methylation was detected by bisulfite conversion and pyrosequencing. MTHFR c.677C>T genotyping was carried out using the TaqMan genotyping assay. Results: LINE-1 methylation level was significantly higher on medial side of cleft lip compared with lateral side (p = 0.001). This difference was not significantly influenced by the case's sex or cleft type. However, MTHFR c.677C>T genotyping revealed that the difference in LINE-1 methylation across cleft lip was restricted to carriers of C allele of MTHFR c.677C>T and was not apparent in TT homozygous cases (p = 0.027). Conclusion: This integrated analysis supports the previous finding of differences in DNA methylation across the two sides of cleft lip and further suggests a possible role of MTHFR c.677C>T genotype in establishing this difference

Rescuing effect of folates on methotrexate cytotoxicity in human trophoblast cells

Objectives: Methotrexate (MTX)is a folate antagonist that is administered in several conditions such as rheumatoid arthritis. Its use may associate with adverse effects presumably originating from folate deficiency. Although MTX side effects could be decreased by folate supplementation, the current guideline on folate administration is not precisely established, which could result in irreversible damage especially in high-risk groups like women in childbearing-age. Thus, this study aimed to investigate the in vitro rescuing effect of different folates including folic acid (FA), 5-methyltetrahydrofolate (MTHF) and folinic acid (5-Formyltetrahydrofolic acid, FTHF) on MTX-treated trophoblast cells. Methods: HTR-8/SVneo cells were stressed with a minimum effective dose of MTX and simultaneously treated with different concentrations of FA, MTHF or FTHF. The rescuing effect was assessed by MTT viability assay. The evaluation was completed by microscopic monitoring, apoptosis assessment and measuring LINE-1 DNA methylation. Results: The MTT viability assay showed no MTX-rescuing effect of FA, but a significant effect of FTHF or MTHF. Microscopic observations supported the results of the viability assay. Accordingly, apoptosis was reduced in MTHF or FTHF treatments, while FA has no effect on the apoptosis induced by MTX. The LINE-1 methylation was not affected by MTX treatment, and not significantly modified in folate supplemented cultures. Conclusions: Despite the general acceptance of administering FA to prevent adverse events of MTX therapy, our findings suggest that FA may not be optimal, and indicate FTHF or MTHF as a better choice. This study on trophoblast cells suggests that FTHF may be the optimal folate, particularly for women in childbearing-age

lncRNA-mediated synovitis in rheumatoid arthritis: A perspective for biomarker development

Long noncoding RNAs (lncRNAs) are a regulatory class of noncoding RNAs with a wide range of activities such as transcriptional and post-transcriptional regulations. Emerging evidence has demonstrated that various lncRNAs contribute to the initiation and progression of Rheumatoid Arthritis (RA) through distinctive mechanisms. The present study reviews the recent findings on lncRNA role in RA development. It focuses on the involvement of different lncRNAs in the main steps of RA pathogenesis including T cell activation, cytokine dysregulation, fibroblast-like synoviocyte (FLS) activation and joint destruction. Besides, it discusses the current findings on RA diagnosis and the potential of lncRNAs as diagnostic, prognostic and predictive biomarkers in Rheumatology clinic

Placental DNA methylation profile as predicting marker for autism spectrum disorder (ASD)

Abstract Autism spectrum disorder (ASD) is a neurodevelopmental disorder that impairs normal brain development and socio-cognitive abilities. The pathogenesis of this condition points out the involvement of genetic and environmental factors during in-utero life. Placenta, as an interface tissue between mother and fetus, provides developing fetus requirements and exposes it to maternal environment as well. Therefore, the alteration of DNA methylation as epigenetic consequence of gene-environmental interaction in the placenta could shed light on ASD pathogenesis. In this study, we reviewed the current findings on placental methylation status and its association with ASD. Differentially methylated regions (DMRs) in ASD-developing placenta were found to be mainly enriched in ASD gene loci affecting synaptogenesis, microtubule dynamics, neurogenesis and neuritogenesis. In addition, non-genic DMRs in ASD-placenta proposes an alternative contributing mechanism for ASD development. Our study highlights the importance of placental DNA methylation signature as a biomarker for ASD prediction

<i>MTHFR</i> c.665C>T and c.1298A>C Polymorphisms in Tailoring Personalized Anti-TNF-α Therapy for Rheumatoid Arthritis

Rheumatoid arthritis (RA) is an inflammatory autoimmune disease with a prevalence of 1%. Currently, RA treatment aims to achieve low disease activity or remission. Failure to achieve this goal causes disease progression with a poor prognosis. When treatment with first-line drugs fails, treatment with tumor necrosis factor-α (TNF-α) inhibitors may be prescribed to which many patients do not respond adequately, making the identification of response markers urgent. This study investigated the association of two RA-related genetic polymorphisms, c.665C>T (historically referred to as C677T) and c.1298A>C, in the MTHFR gene as response markers to an anti-TNF-α therapy. A total of 81 patients were enrolled, 60% of whom responded to the therapy. Analyses showed that both polymorphisms were associated with a response to therapy in an allele dose-dependent manner. The association for c.665C>T was significant for a rare genotype (p = 0.01). However, the observed opposite trend of association for c.1298A>C was not significant. An analysis revealed that c.1298A>C, unlike c.665C>T, was also significantly associated with the drug type (p = 0.032). Our preliminary results showed that the genetic polymorphisms in the MTHFR gene were associated with a response to anti-TNF-α therapy, with a potential significance for the anti-TNF-α drug type. This evidence suggests a role for one-carbon metabolism in anti-TNF-α drug efficacy and contributes to further personalized RA interventions

Seropositivity-Dependent Association between LINE-1 Methylation and Response to Methotrexate Therapy in Early Rheumatoid Arthritis Patients

Methotrexate (MTX) is considered the first choice among disease-modifying anti-rheumatic drugs (DMARDs) for rheumatoid arthritis (RA) treatment. However, response to it varies as approximately 40% of the patients do not respond and would lose the most effective period of treatment time. Therefore, having a predictive biomarker before starting MTX treatment is of utmost importance. Methylation of long interspersed nucleotide element-1 (LINE-1) is generally considered a surrogate marker for global genomic methylation, which has been reported to associate with disease activity after MTX therapy

EphrinA5 regulates cell motility by modulating Snhg15/DNA triplex-dependent targeting of DNMT1 to the Ncam1 promoter

Cell–cell communication is mediated by membrane receptors and their ligands, such as the Eph/ephrin system, orchestrating cell migration during development and in diverse cancer types. Epigenetic mechanisms are key for integrating external “signals”, e.g., from neighboring cells, into the transcriptome in health and disease. Previously, we reported ephrinA5 to trigger transcriptional changes of lncRNAs and protein-coding genes in cerebellar granule cells, a cell model for medulloblastoma. LncRNAs represent important adaptors for epigenetic writers through which they regulate gene expression. Here, we investigate a lncRNA-mediated targeting of DNMT1 to specific gene loci by the combined power of in silico modeling of RNA/DNA interactions and wet lab approaches, in the context of the clinically relevant use case of ephrinA5-dependent regulation of cellular motility of cerebellar granule cells. We provide evidence that Snhg15, a cancer-related lncRNA, recruits DNMT1 to the Ncam1 promoter through RNA/DNA triplex structure formation and the interaction with DNMT1. This mediates DNA methylation-dependent silencing of Ncam1, being abolished by ephrinA5 stimulation-triggered reduction of Snhg15 expression. Hence, we here propose a triple helix recognition mechanism, underlying cell motility regulation via lncRNA-targeted DNA methylation in a clinically relevant context

Prioritization of putatively detrimental variants in euploid miscarriages

Miscarriage is the spontaneous termination of a pregnancy before 24 weeks of gestation. We studied the genome of euploid miscarried embryos from mothers in the range of healthy adult individuals to understand genetic susceptibility to miscarriage not caused by chromosomal aneuploidies. We developed gp , a pipeline that we used to prioritize 439 unique variants in 399 genes, including genes known to be associated with miscarriages. Among the prioritized genes we found STAG2 coding for the cohesin complex subunit, for which inactivation in mouse is lethal, and TLE4 a target of Notch and Wnt, physically interacting with a region on chromosome 9 associated to miscarriages

Prioritization of putatively detrimental variants in euploid miscarriages

none17Miscarriage is the spontaneous termination of a pregnancy before 24 weeks of gestation. We studied the genome of euploid miscarried embryos from mothers in the range of healthy adult individuals to understand genetic susceptibility to miscarriage not caused by chromosomal aneuploidies. We developed GP , a pipeline that we used to prioritize 439 unique variants in 399 genes, including genes known to be associated with miscarriages. Among the prioritized genes we found STAG2 coding for the cohesin complex subunit, for which inactivation in mouse is lethal, and TLE4 a target of Notch and Wnt, physically interacting with a region on chromosome 9 associated to miscarriages.noneBuonaiuto, Silvia; Biase, Immacolata Di; Aleotti, Valentina; Ravaei, Amin; Marino, Adriano De; Damaggio, Gianluca; Chierici, Marco; Pulijala, Madhuri; D’Ambrosio, Palmira; Esposito, Gabriella; Ayub, Qasim; Furlanello, Cesare; Greco, Pantaleo; Capalbo, Antonio; Rubini, Michele; Biase, Sebastiano Di; Colonna, VincenzaBuonaiuto, Silvia; Biase, Immacolata Di; Aleotti, Valentina; Ravaei, Amin; Marino, Adriano De; Damaggio, Gianluca; Chierici, Marco; Pulijala, Madhuri; D’Ambrosio, Palmira; Esposito, Gabriella; Ayub, Qasim; Furlanello, Cesare; Greco, Pantaleo; Capalbo, Antonio; Rubini, Michele; Biase, Sebastiano Di; Colonna, Vincenz