Extracellular vesicles secreted by human aneuploid embryos present a distinct transcriptomic profile and upregulate MUC1 transcription in decidualised endometrial stromal cells.

STUDY QUESTION: Do extracellular vesicles (EVs) secreted by aneuploid human embryos possess a unique transcriptomic profile that elicits a relevant transcriptomic response in decidualized primary endometrial stromal cells (dESCs)? SUMMARY ANSWER: Aneuploid embryo-derived EVs contain transcripts of PPM1J, LINC00561, ANKRD34C, and TMED10 with differential abundance from euploid embryo-derived EVs and induce upregulation of MUC1 transcript in dESCs. WHAT IS KNOWN ALREADY: We have previously reported that IVF embryos secrete EVs that can be internalized by ESCs, conceptualizing that successful implantation to the endometrium is facilitated by EVs. Whether these EVs may additionally serve as biomarkers of ploidy status is unknown. STUDY DESIGN SIZE DURATION: Embryos destined for biopsy for preimplantation genetic testing for aneuploidy (PGT-A) were grown under standard conditions. Spent media (30 µl) were collected from euploid (n = 175) and aneuploid (n = 140) embryos at cleavage (Days 1-3) stage and from euploid (n = 187) and aneuploid (n = 142) embryos at blastocyst (Days 3-5) stage. Media samples from n = 35 cleavage-stage embryos were pooled in order to obtain five euploid and four aneuploid pools. Similarly, media samples from blastocysts were pooled to create one euploid and one aneuploid pool. ESCs were obtained from five women undergoing diagnostic laparoscopy. PARTICIPANTS/MATERIALS SETTING METHODS: EVs were isolated from pools of media by differential centrifugation and EV-RNA sequencing was performed following a single-cell approach that circumvents RNA extraction. ESCs were decidualized (estradiol: 10 nM, progesterone: 1 µM, cAMP: 0.5 mM twice every 48 h) and incubated for 24 h with EVs (50 ng/ml). RNA sequencing was performed on ESCs. MAIN RESULTS AND THE ROLE OF CHANCE: Aneuploid cleavage stage embryos secreted EVs that were less abundant in RNA fragments originating from the genes PPM1J (log2fc = -5.13, P = 0.011), LINC00561 (log2fc = -7.87, P = 0.010), and ANKRD34C (log2fc = -7.30, P = 0.017) and more abundant in TMED10 (log2fc = 1.63, P = 0.025) compared to EVs of euploid embryos. Decidualization per se induced downregulation of MUC1 (log2fc = -0.54, P = 0.0028) in ESCs as a prerequisite for the establishment of receptive endometrium. The expression of MUC1 transcript in decidualized ESCs was significantly increased following treatment with aneuploid compared to euploid embryo-secreted EVs (log2fc = 0.85, P = 0.0201). LARGE SCALE DATA: Raw data have been uploaded to GEO (accession number GSE234338). LIMITATIONS REASONS FOR CAUTION: The findings of the study will require validation utilizing a second cohort of EV samples. WIDER IMPLICATIONS OF THE FINDINGS: The discovery that the transcriptomic profile of EVs secreted from aneuploid cleavage stage embryos differs from that of euploid embryos supports the possibility to develop a non-invasive methodology for PGT-A. The upregulation of MUC1 in dESCs following aneuploid embryo EV treatment proposes a new mechanism underlying implantation failure. STUDY FUNDING/COMPETING INTERESTS: The study was supported by a Marie Sklodowska-Curie Actions fellowship awarded to SM by the European Commission (CERVINO grant agreement ID: 79620) and by a BIRTH research grant from Theramex HQ UK Ltd. The authors have no conflicts of interest to declare.

The relationship between CYP19A1 gene expression in luteinized granulosa cells and follicular estradiol output in women with endometriosis.

Endometriosis was claimed to negatively affect the intrafollicular environment, hindering oocyte competence. Previous studies evaluated expression levels of cytochrome P450 aromatase (CYP19A) in granulosa and cumulus oophorus cells collected from endometriosis women, but results are controversial. To further investigate the intrafollicular environment whose alteration may potentially disturb ovarian steroidogenesis in endometriosis, gene expression of CYP19A and of its upstream enzymes, StAR and 3ßHSD was assessed in luteinized granulosa cells isolated from follicular fluids (FF) collected during Assisted Reproduction Technology (ART) procedures in women with stage III-IV disease and from subjects without the condition. In a subgroup of patients, cumulus oophorus cells (COCs) were also assessed for CYP19A, StAR and 3ßHSD gene expression. No difference in mRNA expression of CYP19A1, StAR and 3ßHSD in both granulosa cells and COCs was observed between the two groups of patients. No significant difference was also found between estradiol FF levels detected in endometriosis patients (median=873, IQR=522-1221 ng/ml)) and control patients (median=878, IQR=609-1137 ng/ml). To gain more insight into the intrafollicular regulation of CYP19A in patients with endometriosis, associations between expression of the analyzed genes, systemic and follicular 17ß-estradiol levels and ART outcomes were assessed. While in the control group, levels of CYP19A1, StAR and 3ßHSD transcripts significantly correlated with follicular estradiol levels (adjusted R² of 0.60), no significant association was detected in affected women (adjusted R² of 0.23). After stratification of the populations based on the presence of the disease, CYP19A1 expression was shown to correlate with the number of oocytes retrieved [ß:- 1.214;95%CI: - 2.085 - (-0.343); p = 0.007] in the control group while this association was not present in patients with endometriosis [ß:- 0.003; 95%CI:- 0.468-0.461; p = 0.988)]. These results do not support data from the literature indicating a reduced aromatase expression in granulosa cells of affected women, but they highlight a potential subtle mechanism affecting the ovulation process in these women.

A Retrospective Observational Real-Word Analysis of the Adherence, Healthcare Resource Consumption and Costs in Patients Treated with Bisoprolol/Perindopril as Single-Pill or Free Combination.

INTRODUCTION: The present real-world analysis aims to compare the drug utilization, hospitalizations and direct healthcare costs related to the use of single-pill combination (SPC) or free-equivalent combination (FEC) of perindopril and bisoprolol (PER/BIS) in a large Italian population. METHODS: This observational retrospective analysis was based on administrative databases covering approximately 7 million subjects across Italy. All adult subjects receiving PER/BIS as SPC or FEC between January 2017-June 2020 were included. Subjects were followed for 1 year after the first prescription of PER/BIS as FEC (± 1 month) or SPC. Before comparing the SPC and FEC cohorts, propensity score matching (PSM) was applied to balance the baseline characteristics. Drug utilization was investigated as adherence (defined by the proportion of days covered, PDC) and persistence (evaluated by Kaplan-Meier curves). Hospitalizations and mean annual direct healthcare costs (due to drug prescriptions, hospitalizations and use of outpatient services) were analyzed during follow-up. RESULTS: The original cohort included 11,440 and 6521 patients taking the SPC and FEC PER/BIS combination, respectively. After PSM, two balanced SPC and FEC cohorts of 4688 patients were obtained (mean age 70 years, approximately 50% male, 24% in secondary prevention). The proportion of adherent patients (PDC ≥ 80%) was higher for those on SPC (45.5%) than those on FEC (38.6%), p < 0.001. The PER/BIS combination was discontinued by 35.8% of patients in the SPC cohort and 41.7% in the FEC cohort (p < 0.001). The SPC cohort had fewer cardiovascular (CV) hospitalizations (5.3%) than the free-combination cohort (7.4%), p < 0.001. Mean annual total healthcare costs were lower in the SPC (1999€) than in the FEC (2359€) cohort (p < 0.001). CONCLUSION: In a real-world setting, patients treated with PER/BIS SPC showed higher adherence, lower risk of drug discontinuation, reduced risk of CV hospitalization, and lower healthcare costs than those on FEC of the same drugs.

Patients with cardiovascular conditions often need to take many pills. This may result in patients not taking their pills as prescribed (i.e., low adherence) and compromise the potential benefits derived from prescription of cardiovascular protective drugs. Simplifying treatment by combining drugs into a single pill can improve adherence and, consequently, patient outcomes. In this analysis using data from real clinical practice, we explored whether using a single pill of perindopril and bisoprolol is associated with higher levels of adherence, lower proportion of patients with hospitalizations and lower economic costs than using the same drugs prescribed as free-equivalent combination in a large sample of the Italian population of approximately 7 million people. We identified two groups of patients taking single pill or free-equivalent combination of perindopril and bisoprolol (4688 patients in each cohort). Over 1-year follow-up, patients taking single pill were more likely to be adherent and were less likely to stop taking their treatment. They also had fewer cardiovascular hospitalizations with shorter hospital admission and had lower healthcare direct costs. In conclusion, simplifying treatment by combining perindopril and bisoprolol in a single pill instead of two may have a positive effect on adherence, outcomes and healthcare costs already after 1 year.

Burden of Illness in Follicular Lymphoma with Multiple Lines of Treatment, Italian RWE Analysis.

This real-world analysis investigated patients with follicular lymphoma in Italy receiving three or more treatment lines (≥3L), focusing on therapeutic pathways with their rebounds on healthcare resource consumptions and costs. Data were retrieved from administrative databases from healthcare entities covering about 13.3 million residents. Adults diagnosed with follicular lymphoma were identified between January 2015 and June 2020, and among them 2434 patients with ≥3L of treatment during the data availability interval (January 2009 to June 2021) were included. Of them, 1318 were in 3L, 494 in 4L and 622 in ≥5L. A relevant proportion of patients (12-32%) switched to a later line within the same calendar year. At 3-year follow-up (median), 34% patients died. Total mean annual expenses were euro 14,508 in the year preceding inclusion and rose to euro 21,081 at 1-year follow-up (on average euro 22,230/patient/year for the whole follow-up), with hospitalization and drug expenses as weightiest cost items. In conclusion, the clinical and economic burden of follicular lymphoma increases along with later treatment lines. The high mortality rates indicate that further efforts are needed to optimize disease management.

Characteristics and Drug Utilization of Patients with Hereditary Angioedema in Italy, a Real-World Analysis.

This real-world analysis investigated the characteristics and treatment patterns of patients with hereditary angioedema (HAE) in Italy using the administrative data of health units across Italy. Patients were identified via exemption code or HAE-specific treatments (thus, all known forms, type I, II and, III, were included). The index date was that of first prescription of HAE treatments within the inclusion period (01/2010-06/2021) or of the date of exemption. The number of HAE patients included was 148 (43.2% male, mean age 43.3 years). Gastrointestinal disorders affected 36.5% patients, hypertension affected 28.4%, hypercholesterolemia affected 11.5%, and depression affected 9.5%. The frequent gastrointestinal involvement was further confirmed by the use of antiemetics and systemic antihistamines that doubled after the index date. Among patients enrolled by treatment (n = 125), n = 105 (84%) were receiving a treatment for acute attacks. This analysis provided insights into the characterization of patients with HAE and their management in Italian clinical practice, suggesting that an unmet therapeutic need could be present for such patients in terms of the clinical burden.

Description of characteristics, management of care and healthcare direct costs of patients with HR+/HER2- early breast cancer in Italy: a real-world study involving administrative and pathological anatomy databases.

BACKGROUND: Healthcare administrative and pathological anatomy data were used to identify Italian patients with early breast cancer (EBC) with HR+/HER2- status at high risk of recurrence, evaluating drug utilization and other healthcare resource use in clinical practice. METHODS: This retrospective analysis, based on 9.4 million of Italian National Health Service beneficiaries, included adult patients with hospitalization discharge diagnosis for EBC in 01/2015-12/2020. Those with HR+/HER2- status were selected; among them, patients that underwent removal of lymph nodes (LN) were analyzed. RESULTS: Of 24,137 patients with EBC and HR+/HER2- status, 3619 patients (15%) had documented LN removal. Overall, 4.7% of HR+/HER2- patients and 9.9% of patients with LN removal experienced distant relapse over a median follow-up of 33.2 months (Q1-Q3: 17.0-50.6). Local relapse occurred in approximately 9.1-9.3% of patients in each group. Among the 1,175 patients with LN removal that had available pathological anatomy data, 399 (34.0%) had pathological high-risk characteristics and 13.3% experienced distant relapse. CONCLUSIONS: One in ten patients with EBC who underwent LN removal experienced a relapse, highlighting the strong need to prevent early recurrence.

Real-world analysis of the economic and therapeutic burden in advanced breast cancer patients in Italy.

BACKGROUND: This real-world analysis evaluated drug utilization focusing on wastage and healthcare costs for treatment of patients with advanced breast cancer (aBC) hormone receptor-positive (HR+)/human epidermal growth factor receptor-2 negative (HER2-) in Italy. METHODS: A retrospective analysis was conducted on administrative data covering about 13.3 million health-assisted individuals. Across January/2017-June/2021, all patients with HR+/HER2-aBC were identified by ≥ 1 prescription for cyclin-dependent kinase 4/6 inhibitors (CDK 4/6i). Cost analysis was performed and updated referring to the prices of November 2021. RESULTS: Overall, 3,647 HR+/HER2-aBC patients were included (2,627 palbociclib treated, 729 ribociclib treated, and 291 abemaciclib treated). After 12 months of follow-up, 35% of palbociclib patients had a dose reduction (on average 8.9 wasted pills/patient), 44.7% of abemaciclib patients had a dose reduction (on average 6.7 wasted pills/patient), 22.1% of ribociclib patients had a dose reduction (no wasted pills). Therapy wastage added up to 528,716€ for palbociclib-treated patients (524€/patient) and 5,738€ in abemaciclib-treated patients (151€/patient). No wastage was attributed to ribociclib. CONCLUSIONS: Dose reduction was associated with drug wastage in palbociclib and abemaciclib-treated patients, but not in ribociclib-treated ones. These findings might be helpful to policy decision-makers who, for healthcare strategies implementation, among several variables should consider the possible restraining of drug wastage.

Epidemiology, Patients' Journey and Healthcare Costs in Early-Stage Non-Small-Cell Lung Carcinoma: A Real-World Evidence Analysis in Italy.

This real-world analysis aims to estimate the epidemiology and economic burden related to early-stage non-small-cell lung carcinoma (eNSCLC) in the clinical practice Italian setting. An observational analysis was performed using administrative databases linked to pathological anatomy data, covering around 2.5 mln health-assisted individuals. From 2015 to mid-2021, eNSCLC patients staged II-IIIA treated with chemotherapy after surgery were included. Patients were stratified into those presenting loco-regional or metastatic recurrence during follow-up and annualized healthcare direct costs covered by the Italian National Health System (INHS) were estimated. In 2019-2020, the prevalence of eNSCLC was 104.3-117.1/million health-assisted subjects, and the annual incidence was 38.6-30.3/million. Data projected to the Italian population estimated 6206 (2019) and 6967 (2020) prevalent and 2297 (2019) and 1803 (2020) incident cases. Overall, 458 eNSCLC patients were included. Of them, 52.4% of patients had a recurrence (5% loco-regional-recurrence, 47.4% metastatic-recurrence). Healthcare total direct costs/patient averaged EUR 23,607, in particular, in the first year after recurrence, costs averaged EUR 22,493 and EUR 29,337 in loco-regional and metastatic-recurrence patients, respectively. This analysis showed that about one-half of eNSCLC patients stage II-IIIA experience a recurrence, and in recurrence patients, total direct costs were almost two-fold those of no-recurrence patients. These data highlighted an unmet clinical need, as the therapeutic optimization of patients at early stages.

The Economic Burden of Chronic Myeloid Leukemia in Patients with Later Lines: Findings from a Real-World Analysis in Italy.

INTRODUCTION: Chronic myeloid leukemia (CML) is a hematopoietic myeloproliferative disorder that accounts for 20% of all leukemias of adults. The introduction of tyrosine kinase inhibitors (TKIs) (imatinib, bosutinib, dasatinib, nilotinib, ponatinib) has yielded significant benefits for patients with CML in terms of survival and quality of life. This real-world analysis evaluated the economic burden for managing patients with CML in 2nd or ≥ 3rd TKI lines in Italian settings of clinical practice. METHODS: A retrospective observational analysis was performed exploiting the administrative databases of a sample of entities covering around 15 million inhabitants. From 2015 to 2018, the study included adult patients with at least one prescription for TKIs, (and for some TKI with at least one hospitalization discharge diagnosis for CML, or at least one prescription for BCR-ABL examination). The index date was the first TKI prescription. Healthcare resource consumption and costs for patients with CML in 2nd and ≥ 3rd line treatment with TKIs were analyzed for drug prescriptions, hospitalizations, specialist visits, and diagnostic services. RESULTS: In total 635 patients were included, 491 in 2nd line and 144 in 3rd line with TKIs. Dasatinib was the most frequently prescribed drug in 2nd line (28.9%) and imatinib in later lines (26.4%). With progressing lines of treatment, healthcare consumption showed a trend towards increased non-TKI prescriptions per patient (8 for 2nd line and 9.7 for ≥ 3rd line). The management of patients with CML in later lines resulted in increased overall healthcare burden, with hospitalizations accounting for about half of total expenditure, whatever the treatment line and type of TKI. CONCLUSIONS: This analysis in Italian real-life clinical practice reported economic expenditure for patients with CML in 2nd or ≥ 3rd lines with TKIs, mostly burdened by hospitalizations. Such clinical complexity suggests that further efforts are needed to improve the therapeutic management of later lines of CML.

The Epidemiology, Treatment Patterns and Economic Burden of Different Phenotypes of Multiple Sclerosis in Italy: Relapsing-Remitting Multiple Sclerosis and Secondary Progressive Multiple Sclerosis.

Purpose: A retrospective analysis of real-world data was performed to assess the epidemiology and economic burden of multiple sclerosis (MS), relapsing-remitting MS (RRMS), and secondary-progressive MS (SPMS) in Italy. Patients and Methods: An observational study on administrative databases from a sample of Italian entities was carried-out. Between 01/2010-12/2017, patients with ≥1 MS diagnosis code (ICD-9-CM:340 and/or exemption code:046) and/or ≥1 disease-modifying therapies (DMTs) prescription, were included. Among MS-cohort, SPMS patients were identified by ≥2 hospitalizations or by ≥2 drug prescriptions related to MS progression. MS patients not fulfilling SPMS criteria were included as RRMS. Mean annual healthcare costs were reported during follow-up and stratified by DMT treatment/untreatment. Results: Overall, 9543 MS patients were included; 8397 with RRMS and 1146 with SPMS. Estimated prevalence of MS was 141.6/100,000 inhabitants (RRMS 124.4/100,000 and SPMS 17.2/100,000). Mean annual cost for untreated and treated patient was respectively: €3638 and €11796 (MS-cohort), €3183 and €11486 (RRMS-cohort), €6317 and €15511 (SPMS-cohort). The first-line DMT treatment duration averaged 27.4 ± 22.8 months; the mean cost was 19004€ for the whole period. The second-line DMT treatment lasted on average 31.1 ± 24.5 months; the mean cost was 47293€ for the whole period. Conclusion: This study provided insights into the MS epidemiology in Italy and its economic burden. Healthcare costs associated with MS management were mainly driven by DMTs expenditure. A trend of higher healthcare-resource consumption was observed among SPMS-cohort.

Evaluation of the Therapeutic Pattern and Pharmaco-Utilization in Hypercholesterolemic Patients Treated with Statins: A Retrospective Study on Italian Real-World Data.

Purpose: The study aimed to analyze, in hypercholesterolemic patients under statin medication, patient characteristics and their lipid profile at baseline, the therapeutic pathway, and the pharmaco-utilization, using real-world data in Italy. Patients and Methods: A retrospective study was conducted using administrative databases of a sample of entities covering 6.5 million health-assisted individuals. Between January 2010 and June 2019, patients with non-familial hypercholesterolemia (nFH) were identified by 1) ≥1 low-density lipoprotein cholesterol (LDL-C) measurement (LDL-C assessment date was the index-date) and 2) statin prescription during 6 months before the index-date (pharmaco-utilization period). FH patients were defined by LDL-C evaluation, statin treatment during the pharmaco-utilization period, and a score ≥6 according to the Dutch Lipid Clinic Network criteria. nFH patients were divided into four exclusive cohorts based on CV-risk class: 1) with previous CV disease (CVD); 2) with diabetes mellitus; 3) with mixed-dyslipidemia diagnosis; 4) in primary-prevention. Based on LDL-C index values, patient was defined with LDL-C "controlled" if its levels were ≤70mg/dl (CVD), ≤100mg/dl (diabetes, FH), ≤130mg/dl (mixed-dyslipidemia, primary-prevention). Results: Overall 164,161 nFH patients were included (mean age 72 years, 51% male); of these, 46,782 (28.5%) were CVD (mean age 74 years, 66% male), 34,803 (21.2%) were diabetic (mean age 72 years, 51% male), 1617 (1%) were with mixed-dyslipidemia (mean age 71 years, 48% male) and 80,959 (49.3%) were in primary-prevention (mean age 71 years, 42% male). The proportion of nFH patients with controlled LDL-C was 41.2% for CVD, 73.6% for diabetic, 80.7% for mixed-dyslipidemia, and 79.5% for primary-prevention patients; 49% of nFH patients were adherent to therapy. Overall, 1287 FH patients (mean age 64 years, 42% male) were included; in 39.2% of the patients, LDL-C was controlled, and 44% of the patients were adherent to therapy. Conclusion: The results of this study highlighted non-optimal therapeutic management of hypercholesterolemic patients in Italian clinical practice, with a notable quote of patients non-adherent to therapy.

Real-World Analysis of the Therapeutic Management and Disease Burden in Chronic Myeloid Leukemia Patients with Later Lines in Italy.

Real world data are becoming a crucial tool to understand how cancer is treated in routine daily practice. This real-world analysis aims to describe the characteristics of patients with CML in 2nd or ≥3rd tyrosine kinase inhibitors (TKI) lines of therapy, to evaluate their treatment sequence and utilization in settings of Italian clinical practice in Italy. A retrospective analysis was performed using an administrative databases covering around 15.3 million cases. All adult patients prescribed with TKI as 2nd or ≥3rd lines (L) of therapy for CML during January 2015-December 2018 were included. A total of 491 patients in 2nd and 144 in ≥3rd L was included. In both cohorts, hypertension was the most reported comorbidity, followed by metabolic and blood count alterations. In each calendar inclusion year, an increment of 97.6% was observed in the number of patients treated in ≥3rd L. In the 2nd L cohort, 18.7% had a switch to 3rd L, while 26.4% of ≥3rd L patients switched to a subsequent line. Around 40% in both lines discontinued their treatment after a median time of 5.5 (2nd L) and 4.3 (≥3rd L) years. The results provided insights into CML management clinical practice, indicating a heavy disease burden for patients in later lines that showed an increasing complex management, and suggest that a need for novel treatment strategies might exists.

Uterine fluid cytokine/chemokine levels of women undergoing ART with and without oral Vitamin D supplementation.

STUDY QUESTION: Is oral Vitamin D supplementation able to modify the intrauterine milieu in terms of cytokine/chemokine pattern? SUMMARY ANSWER: No significant differences were detected in cytokine and chemokine levels in endometrial secretions between patients undergoing ART with or without Vitamin D supplementation. WHAT IS KNOWN ALREADY: Cytokines and chemokines secreted into the intrauterine environment are fundamental for the molecular crosstalk between the endometrium and the preimplantation embryo. Whether Vitamin D can regulate these mediators in the endometrial environment is still unclear. STUDY DESIGN SIZE DURATION: This study was an analysis of a secondary outcome from the Supplementation of Vitamin D and Reproductive Outcomes-SUNDRO-clinical trial, a multicenter randomized double-blinded trial designed to explore the effects of Vitamin D replacement in women with Vitamin D levels below 30 ng/ml undergoing autologous ART cycles. Uterine fluid samples were collected from both patients supplemented with Vitamin D (n = 17) and from the placebo group (n = 32). PARTICIPANTS/MATERIALS SETTING METHODS: Based on cutoff points for Vitamin D insufficiency (20-29.9 ng/ml) or deficiency (<20 ng/ml), 67% of patients in the study were insufficient, and 33% deficient, in Vitamin D, although they were considered together for the analysis. Women received a single dose of 600 000 IU 25-hydroxyvitamin D or placebo from 2 to 12 weeks before oocyte retrieval. Inclusion criteria were female age 18-39 years, with a BMI between 18 and 25 kg/m2. Serum 25-hydroxyvitamin D was assessed at the time of hCG administration. Uterine fluid samples were collected during the secretory phase of the menstrual cycle preceding oocyte retrieval. The quantitative determination of 27 cytokines in endometrial secretion samples was performed by using a multiplex immunoassay. MAIN RESULTS AND THE ROLE OF CHANCE: Uterine fluid samples were collected after a median (range) of 21 (12-41) days after the oral Vitamin D supplementation. Both the supplemented and placebo groups had Vitamin D serum levels below 30 ng/ml at baseline/time of randomization ((median 23.4 ng/ml (interquartile range 19.5-28.4) and 23.4 ng/ml (17.8-25.9), respectively). At the time of hCG administration, serum Vitamin D in supplemented subjects was significantly raised compared to the placebo group ((median 52.9 ng/ml (interquartile range 40.7-64.1) and 24.6 ng/ml (19.3-29.2), respectively, P < 0.001). Our data revealed no significant differences in uterine fluid cytokine/chemokine composition of Vitamin D-supplemented women compared with the placebo group. This finding remained when the concentrations of all mediators studied were normalized to total protein. In a further analysis, no significant differences were found in the content of cytokines/chemokines in uterine fluid from women who conceived (n = 19) compared with the nonpregnant group (n = 30). LIMITATIONS REASONS FOR CAUTION: Using a randomized study design (a single dose of 600 000 IU 25-hydroxyvitamin D versus placebo), we found no significant differences between groups. However, we cannot exclude that any benefit of Vitamin D supplementation may be specific for some subgroups of patients, such as those with an imbalance of T-helper 1 and T-helper 2 cell populations. The uterine secretions were collected during the menstrual cycle that preceded oocyte retrieval; therefore, it is possible the uterine fluid collection and analysis in the same cycle of the embryo transfer might have resulted in different conclusions. Moreover, the small sample size could limit the power of the study. WIDER IMPLICATIONS OF THE FINDINGS: Our analysis of the uterine secretome profiling failed to show any significant difference in endometrial cytokine/chemokine patterns between women with oral Vitamin D supplementation and the placebo group. Vitamin D may act on the uterine environment through a different mechanism. STUDY FUNDING/COMPETING INTERESTS: The study was funded by the Italian Ministry of Health following peer review in the competitive 'Bando di Ricerca Finalizzata e Giovani Ricercatori 2013' with reference code RF-2013-02358757. The authors declare that they have no conflict of interest. TRIAL REGISTRATION NUMBER: EudraCT registration number: 2015-004233-27.

Direct Healthcare Costs by Level of Adherence of a Real-World Population of Statin Users in Italy.

Purpose: This real-world study investigates the direct healthcare costs from the perspective of the Italian Healthcare National Service of experienced statin users according to their level of adherence to therapy and to their cardiovascular (CV) profile in Italian settings of outpatients clinical practice. Patients and Methods: A retrospective observational analysis was performed based on administrative databases covering approximately 6 million health-assisted individuals. Adult patients with statins prescription between January 2014 and December 2016 were screened, and first prescription within this period was the index date. Follow-up lasted 1 year after index date. Only patients receiving statins prior index date (experienced statin users) were included and distributed in clusters based on their CV profile. Adherence was calculated during follow-up as proportion of days covered (PDC) and classified in low adherence (PDC<40%), partial adherence (PDC=40-79%) and adherence (PDC≥80%). Mean direct healthcare costs of drugs, hospitalizations, and outpatient services were evaluated during follow-up. Results: A total of 436,623 experienced statin users were included and distributed as follows: 5.5% in the previous CV events, 22.6% in diabetes, 55.7% in CV treatments and 16.2% in the no comorbidity cluster. Total mean annual cost/patient decreased from low adherent to adherent patients from €4826 to €3497 in previous CV events, from €2815 to €2360 in diabetes cluster, from €2077 to €1863 for patients with CV treatments. Same trend was reported for the cost item related to hospitalizations, which was the major determinant of the total costs. In previous CV event cluster, adherence was associated to a saving of €879 on total costs. Conclusion: The study highlighted a decrease in overall mean costs as adherence levels increase, particularly for patients with previous CV events, showing how improving adherence could be associated to cost savings and suggesting suited strategy based on CV profile should be undertaken for adherence optimization.

Cellular and Molecular Nature of Fragmentation of Human Embryos.

Embryo fragmentation represents a phenomenon generally characterized by the presence of membrane-bound extracellular cytoplasm into the perivitelline space. Recent evidence supports the cellular and molecular heterogeneity of embryo fragments. In this narrative review, we described the different embryo fragment-like cellular structures in their morphology, molecular content, and supposed function and have reported the proposed theories on their origin over the years. We identified articles related to characterization of embryo fragmentation with a specific literature search string. The occurrence of embryo fragmentation has been related to various mechanisms, of which the most studied are apoptotic cell death, membrane compartmentalization of altered DNA, cytoskeletal disorders, and vesicle formation. These phenomena are thought to result in the extrusion of entire blastomeres, release of apoptotic bodies and other vesicles, and micronuclei formation. Different patterns of fragmentation may have different etiologies and effects on embryo competence. Removal of fragments from the embryo before embryo transfer with the aim to improve implantation potential should be reconsidered on the basis of the present observations.

Analysis of the prevalence of ankylosing spondylitis and treatment patterns and drug utilization among affected patients: an Italian real-world study.

OBJECTIVES: To evaluate the prevalence of ankylosing spondylitis (AS) and associated treatment patterns and drug utilization in real-world clinical practice in Italy. METHODS: This observational study used data from administrative databases of selected Italian entities and included patients with AS diagnosis identified by ICD-9-CM and/or exemption codes. Patients without biologic disease-modifying antirheumatic drug (bDMARD) prescriptions prior to inclusion were defined bio-naïve. A cross-sectional analysis identified treatment patterns from 2016 to 2018. A longitudinal analysis investigated bDMARD utilization (persistence, switch, discontinuing treatment) among bio-naïve patients, with 2014 and 2017 as inclusion periods. Follow-up extended from first bDMARD prescription (index date) to end of data availability. RESULTS: The prevalence of AS was 75.5 per 100,000 (88.3 per 100,000 in adults) in 2018. In total, 5,942 AS patients were identified in 2016, 6,554 in 2017, and 7,146 in 2018. bDMARDs were prescribed to 21.4% (2016), 22.0% (2017), and 16.9% (2018) of the patients. Among the 349 patients included in 2014, 9.5% switched therapies, 13.8% discontinued treatment. In 2017, 12.2% of the 262 patients included switched therapies, 27.1% discontinued treatment. CONCLUSIONS: This study provided an up-to-date prevalence of AS diagnosis in Italy. Bio-naïve patients showed an increasing tendency toward switch of therapy and discontinuation.

Treatment Patterns and Pharmacoutilization in Patients Affected by Psoriasis: An Observational Study in an Italian Real-World Setting.

BACKGROUND: Real-world data can inform the use of biologics for psoriasis (PSO). OBJECTIVE: The aim was to evaluate treatment patterns and analyze pharmacoutilization in PSO patients in a real-world Italian setting, with a focus on the biologics most recently introduced. METHODS: An observational study based on administrative databases was conducted. Patients were included based on PSO diagnosis identified by either discharge diagnosis or exemption code or prescription of anti-psoriatic topical drugs (proxy of diagnosis). To describe patient characteristics and treatment patterns using the most up-to-date data, two different approaches were used: a cross-sectional study performed during 2016-2018, and a longitudinal study conducted with patients who received their first biological/targeted synthetic drugs (naïve patients) in 2014 and 2017 (the inclusion periods). RESULTS: During 2016-2018, the number of prevalent patients diagnosed with PSO was 194,054 (2016), 210,830 (2017), and 225,171 (2018). The percentage of patients receiving biologics or targeted synthetic agents ranged from 1.5 to 2.1%. Among them, naïve patients receiving interleukin (IL) inhibitors increased from 37.5% (2016) to 69.4% (2018), while those receiving anti-tumor necrosis factor (anti-TNF) decreased from 62.5% (2016) to 30.6% (2018). The longitudinal analysis included 894 and 1218 naïve patients in 2014 and 2017, respectively, of whom 7.2% (2014) and 6.9% (2017) switched therapy after a mean of 7.1 (2014) and 6.9 (2017) months. Overall, 259 patients were prescribed ixekizumab starting in 2017, of whom 73% were naïve. Ixekizumab was prescribed as monotherapy to 52.5%. CONCLUSIONS: The proportion of patients receiving biologics appeared constant over the years, with an increasing number of naïve patients being prescribed IL-17 inhibitors. Ixekizumab patients were mostly naïve.

Association between hyperkalemia, RAASi non-adherence and outcomes in chronic kidney disease.

BACKGROUND: Hyperkalemia is relatively frequent in CKD patients treated with renin-angiotensin-aldosterone-system inhibitors (RAASi). AIM: The aim of the present study was to estimate the increased risk of cardiovascular events and mortality due to sub-optimal adherence to RAASi in CKD patients with hyperkalemia. METHODS: An observational retrospective cohort study was conducted, based on administrative and laboratory databases of five Local Health Units. Adult patients discharged from the hospital with a diagnosis of CKD, who were prescribed RAASi between January 2010 and December 2017, were included. We evaluated the appearance of documented episodes of hyperkalemia, RAASi therapy adherence and the effects of these two variables on cardiovascular events, death and dialysis inception for study patients. RESULTS: Of the 9241 selected patients, 4451 met all the criteria for study inclusion. Among them, 1071 had at least one documented episode of hyperkalemia, while 3380 did not. After propensity score matching based on several variables we obtained 2 groups of patients. The appearance of hyperkalemia caused treatment discontinuation in 21.8% of patients previously on RAASi therapy, and sub-optimal adherence (proportion of days covered < 80%) in 33.6% of them. Non-adherence to RAASi therapy among hyperkalemia patients was associated with a higher risk of cardiovascular events (hazard ratio [HR] 1.45, confidence interval [CI] 1.02-2.08; p < 0.05). Moreover, in non-adherent hyperkalemia patients, the risk of death increased by 126% (HR 2.26, CI 1.62-3.15; p < 0.001) compared with adherent patients. CONCLUSIONS: In a large cohort of CKD patients treated with RAASi, we observed that following hyperkalemia onset, non-adherence to RAASi medication can result in an increased risk of cardiovascular events and death.

Association between short-term exposure to particulate matter air pollution and outcomes of assisted reproduction technology in Lombardy, Italy: A retrospective cohort study.

The harmful effect of polluted air on spontaneous fertility has been consistently reported. However, the specific pollutants involved in determining this effect are still to be clarified. The study of Assisted Reproductive Technology (ART) populations is particularly helpful in this context since it allows to monitor the key events of the reproductive process. We analyzed the medical records of 2122 patients who underwent fresh or frozen ART cycles during 2014-2017 in the Lombardy region, north-west Italy. Each subject was assigned the daily PM10 estimates concentration, at the municipality of residence, during the induction of multiple follicular growth. A multivariable linear regression model with a repeated-measures design was used to estimate the association between short-term exposure to PM10 and ART outcomes, A reduction in the number of retrieved oocytes in association with 10 µg/m3 increment of the pollutant estimated at 13-14 days before oocyte retrieval (Day 0) and a decrease in the percentage of metaphase II oocytes for 1-week and 2-weeks mean exposure before day 0 were observed. For cumulative pregnancies, a significant lag time change effect for PM10 exposure (Day -9-0) has been detected, by means of multivariable logistic regression models. An increase in PM10 exposure was associated with a decrease in clinical and ongoing pregnancies while a decrease in PM10 exposure was associated with a significant increase in pregnancy rates. In a population living in a highly polluted area in Italy, we added suggestive evidence of a negative association between ART outcomes and PM10 exposure after controlling for known risk factors for ART success rate.