Fertility specialists' views, behavior, and attitudes towards the use of endometrial scratching in Italy.

BACKGROUND: Endometrial scratching (ES) or injury is intentional damage to the endometrium performed to improve reproductive outcomes for infertile women desiring pregnancy. Moreover, recent systematic reviews with meta-analyses and randomized controlled trials demonstrated that ES is not effective, data on the safety are limited, and it should not be recommended in clinical practice. The aim of the current study was to assess the view and behavior towards ES among fertility specialists throughout infertility centers in Italy, and the relationship between these views and the attitudes towards the use of ES as an add-on in their commercial setting. METHODS: Online survey among infertility centers, affiliated to Italian Society of Human Reproduction (SIRU), was performed using a detailed questionnaire including 45 questions with the possibility to give "closed" multi-choice answers for 41 items and "open" answers for 4 items. Online data from the websites of the infertility centers resulting in affiliation with the specialists were also recorded and analyzed. The quality of information about ES given on infertility centers websites was assessed using a scoring matrix including 10 specific questions (scored from 0 to 2 points), and the possible scores ranged from 0 to 13 points ('excellent' if the score was 9 points or more, 'moderate' if the score was between 5 and 8, and 'poor' if it was 4 points or less). RESULTS: The response rate was of 60.6% (43 questionnaires / 71 infertility SIRU-affiliated centers). All included questionnaires were completed in their entirety. Most physicians (~ 70%) reported to offer ES to less than 10% of their patients. The procedure is mainly performed in the secretory phase (69.2%) using pipelle (61.5%), and usually in medical ambulatory (56.4%) before IVF cycles to improve implantation (71.8%) without drugs administration (e.g., pain drugs, antibiotics, anti-hemorrhagics, or others) before (76.8%) or after (64.1%) the procedure. Only a little proportion of infertility centers included in the analysis proposes formally the ES as an add-on procedure (9.3%), even if, when proposed, the full description of the indications, efficacy, safety, and costs is never addressed. However, the overall information quality of the websites was generally "poor" ranging from 3 to 8 and having a low total score (4.7 ± 1.6; mean ± standard deviation). CONCLUSIONS: In Italy, ES is a procedure still performed among fertility specialists for improving the implantation rate in IVF patients. Moreover, they have a poor attitude in proposing ES as an add-on in the commercial setting.

The true panel of cystic fibrosis mutations in the Sicilian population.

BACKGROUND: The aim was to establish the true risk of having an affected child with Cystic Fibrosis (CF) in the Sicilian infertile population. METHODS: A longitudinal CFTR screening of 1279 Sicilian infertile patients for all CFTR mutations sequencing the entire gene by Next Generation Sequencing (NGS) was performed from patient's blood. RESULTS: One patient out of 16 was a carrier of a CFTR mutation. Twenty-four mutations were found. Theoretically one couple out of 256 was at risk of CF transmission. CONCLUSIONS: The risk of CF transmission is unexpectedly high in Sicily and with a high heterogeneity. Sequencing an entire and long gene such as CFTR makes accessible the true panel of mutations in a specific population and helps better to understand the true risk of having an affected child.

Universal strategy for preimplantation genetic testing for cystic fibrosis based on next generation sequencing.

PURPOSE: We developed and applied a universal strategy for preimplantation genetic testing for all cystic fibrosis gene mutations (PGT-CF) based on next-generation sequencing (NGS). METHODS: A molecular protocol was designed to diagnose all CF mutations at preimplantation stage. The detection of CF mutations was performed by direct gene sequencing and linkage strategy testing 38 specific SNPs located upstream and inside the gene for PGT-CF. Seventeen couples at risk of CF transmission decided to undergo PGT-CF. Trophectoderm cell biopsies were performed on day 5-6 blastocysts. PGT for aneuploidy (PGT-A) was performed from the same samples. Tested embryos were transferred on further natural cycles. RESULTS: PGT was performed on 109 embryos. Fifteen CF mutations were tested. PGT-CF and PGT-A were conclusive for respectively 92.7% and 95.3% of the samples. A mean of 24.1 SNPs was informative per couple. After a single embryo transfer on natural cycle, 81.3% of the transferred tested embryos were implanted. CONCLUSIONS: The present protocol based on the entire CFTR gene together with informative SNPs outside and inside the gene can be applied to diagnose all CF mutations at preimplantation stage.

Universal strategy for preimplantation genetic testing for cystic fibrosis based on next-generation sequencing.

PURPOSE: We developed and applied a universal strategy for preimplantation genetic testing for all cystic fibrosis gene mutations (PGT-CF) based on next-generation sequencing (NGS). METHODS: A molecular protocol was designed to diagnose all CF mutations at preimplantation stage. The detection of CF mutations was performed by direct gene sequencing and linkage strategy testing 38 specific SNPs located upstream and inside the gene for PGT-CF. Seventeen couples at risk of CF transmission decided to undergo PGT-CF. Trophectoderm cell biopsies were performed on days 5-6 blastocysts. PGT for aneuploidy (PGT-A) was performed from the same samples. Tested embryos were transferred on further natural cycles. RESULTS: PGT was performed on 109 embryos. Fifteen CF mutations were tested. PGT-CF and PGT-A were conclusive for, respectively, 92.7% and 95.3% of the samples. A mean of 24.1 SNPs was informative per couple. After single embryo transfer on natural cycle, 81.3% of the transferred tested embryos implanted. CONCLUSIONS: The present protocol based on the entire CFTR gene sequencing together with informative SNPs outside and inside the gene can be applied to diagnose all CF mutations at preimplantation stage.

The accumulation of vitrified oocytes is a strategy to increase the number of euploid available blastocysts for transfer after preimplantation genetic testing.

PURPOSE: In a preimplantation genetic diagnosis for aneuploidy (PGD-A) program, the more embryos available for biopsy, consequently increases the chances of obtaining euploid embryos to transfer. The aim was to increase the number of viable euploid blastocysts in patients undergoing PGD-A using fresh oocytes together with previously accumulated vitrified oocytes. METHODS: Sixty-nine patients with normal ovarian reserve underwent PGD-A for repeated implantation failure or recurrent pregnancy loss indication. After several cycles of ovarian stimulation, 591 accumulated vitrified oocytes and 463 fresh oocytes were micro-injected with the same partner's semen sample. PGD-A was completed on 134 blastocysts from vitrified/warmed oocytes and 130 blastocysts from fresh oocytes. RESULTS: A mean of 9.6% euploid blastocyst per micro-injected vitrified/warmed oocytes and 11.4% euploid blastocyst per micro-injected fresh oocyte were obtained (p > 0.05). The euploidy and aneuploidy rates were comparable in blastocysts obtained from micro-injected vitrified/warmed oocytes and fresh oocytes (42.5 versus 40.8% and 57.5 versus 59.2%, p > 0.05). Implantation rates of euploid blastocysts were comparable between the two sources of oocytes (56.0% from vitrified/warmed oocytes versus 60.9% from fresh oocytes, p > 0.05). CONCLUSIONS: Oocyte vitrification and warming do not generate aneuploidy in blastocysts. The number of viable euploid embryos for transfer can be increased by using accumulated vitrified oocytes together with fresh oocytes in ICSI. TRIAL REGISTRATION: NCT02820415 ClinicalTrials.gov.

Different Tissue-Derived Stem Cells: A Comparison of Neural Differentiation Capability.

BACKGROUND: Stem cells are capable of self-renewal and differentiation into a wide range of cell types with multiple clinical and therapeutic applications. Stem cells are providing hope for many diseases that currently lack effective therapeutic methods, including strokes, Huntington's disease, Alzheimer's and Parkinson's disease. However, the paucity of suitable cell types for cell replacement therapy in patients suffering from neurological disorders has hampered the development of this promising therapeutic approach. AIM: The innovative aspect of this study has been to evaluate the neural differentiation capability of different tissue-derived stem cells coming from different tissue sources such as bone marrow, umbilical cord blood, human endometrium and amniotic fluid, cultured under the same supplemented media neuro-transcription factor conditions, testing the expression of neural markers such as GFAP, Nestin and Neurofilaments using the immunofluorescence staining assay and some typical clusters of differentiation such as CD34, CD90, CD105 and CD133 by using the cytofluorimetric test assay. RESULTS: Amniotic fluid derived stem cells showed a more primitive phenotype compared to the differentiating potential demonstrated by the other stem cell sources, representing a realistic possibility in the field of regenerative cell therapy suitable for neurodegenerative diseases.