What has to be pointed out in unexplained recurrent pregnancy loss research in the unsolved fields: Lessons from clinic. An Italian RPL Unit experience

Recurrent pregnancy loss (RPL) is a controversial field both in research and clinical approaches. Despite the most recent guidelines (ESHRE 2017), an agreement in diagnostic work-up to apply in these patients, as well as in management and treatment, has not been reached, especially in unexplained RPL (uRPL). This is due to the lack of a strong evidence-based level in this field, since the discrepancies among the different RPL research groups, in terms of definition, etiological factors and management cannot lead to organize all the results in systematic reviews. Therefore, common shared cornerstones are required to homogenize research parameters, since the right interplay between clinical management and experimental approaches could lead to contribute in the development of stronger evidences. In this review, we highlight what has to be pointed out in RPL debated subfields and how the experimental approach is necessary to overhaul discrepancies in clinical management. The experience of our RPL Unit has been reported to show how research experience could contribute in modifying the clinical approach

Transoral robotic total laryngectomy

International audienc

Three-hour analysis of non-invasive foetal sex determination: application of Plexor chemistry

Contesto: La conoscenza del singolo "status" genetico in epoca prenatale è particolarmente rilevante nel caso di storia familiare positiva per le malattie genetiche, in età materna avanzata e in screening generale per anomalie fetali. In questo contesto, qui, riportiamo un saggio molecolare innovativo che utilizza il DNA fetale acellulare (cffDNA) come fonte per la diagnosi precoce e rapida del sesso fetale. Lo studio ha coinvolto 132 donne in gravidanza nei primi 3 mesi di gravidanza, che hanno accettato di dare un campione di sangue. Tutti i campioni raccolti sono stati immediatamente sottoposti alla separazione del plasma, che è stato utilizzato per l'estrazione del cffDNA. Successivamente, il cffDNA estratto è stato analizzato con un metodo PCR quantitativo (qPCR) basato su Plexor-HY chimico, che è in grado di identificare simultaneamente, quantificare e discriminare il DNA autosomico dal DNA legato al sesso. Risultati: In generale, il dosaggio Plexor-HY ha dimostrato di essere sensibile e specifico per la determinazione del DNA lowtemplate, come il cffDNA. Infatti, il saggio Plexor-HY è stato eseguito con successo in tutti i campioni, identificando 70 maschi e 62 femmine. Come il sesso fetale può essere fornito in 120 min solo utilizzando un campione di sangue materno come sorgente cffDNA, il saggio rappresenta un metodo prenatale molto veloce, sicuro e non invasivo. Conclusioni: La possibilità di determinare il sesso del feto nella vita prenatale precoce consente la applicazione del nostro test come test di screening utile per i soggetti e le famiglie a rischio di patologie legate al sesso. Inoltre, la conoscenza anticipata del sesso del feto può essere di grande aiuto anche per lo specialista, che potrebbe prontamente consigliare i pazienti sui rischi fetali di ereditare disturbi legati al sesso e l'utilità clinica di eseguire una diagnosi prenatale invasiva.Background: The knowledge of the individual genetic “status” in the prenatal era is particularly relevant in the case of positive family history for genetic diseases, in advanced maternal age and in the general screening for foetal abnormalities. In this context, here, we report an innovative molecular assay which utilizes the cell-free foetal DNA (cffDNA) as a source for the early and fast detection of the foetal sex. The study involved 132 pregnant women in their first 3 months of pregnancy, who agreed to give a blood sample. All the collected samples were immediately subjected to the separation of the plasma, which was utilized for the extraction of the cffDNA. Successively, the extracted cffDNA was analysed by a quantitative PCR (qPCR) method based on Plexor-HY chemistry, which is able to simultaneously identify, quantify and discriminate the autosomal DNA from the sex-linked DNA. Results: Overall, the Plexor-HY assay demonstrated to be sensitive and specific for the determination of lowtemplate DNA, such as the cffDNA. In fact, the Plexor-HY assay has been successfully performed in all the samples, identifying 70 males and 62 females. As the foetal sex can be provided in 120 min just by utilizing a maternal blood sample as cffDNA source, the assay represents a very fast, safe and non-invasive prenatal method. Conclusions: The possibility of determining the foetal sex in the early prenatal life consents the application of our assay as a helpful screening test for subjects and families at risk of sex-linked disorders. Moreover, the early knowledge of the foetal sex may be of great help even for the specialist, who might promptly advise the patients concerning the foetal risk of inheriting sex-linked disorders and the clinical utility of performing an invasive prenatal diagnosis

Transabdominal coelocentesis as early source of fetal DNA for chromosomal and molecular diagnosis

This study reports a comparative analysis between results of transabdominal coelocentesis and traditional invasive procedure in order to assess the usefulness of coelocentesis as a source of fetal DNA for molecular and chromosomal analysis. A number of 28 women were included in the study. A successful sampling of coelomic fluid was obtained in 25 women by transabdominal procedure. A positive amplification of DNA with QF-PCR techniques was obtained in 90% of cases, while 10% of cases failed to reveal interpretable results. Although all samples were cultured, the growth rate was not sufficient to determine karyotypes within 2 weeks. Five samples were selected to be analyzed by array-based comparative genomic hybridization (a-CGH) but the interpretation of these results was difficult and ambiguous. Our results suggest that transabdominal coelocentesis is suitable for the detection of single DNA variation and for QF-PCR analysis, while further experiments are needed to develop optimized protocols for traditional karyotyping and array-analysis