Transcriptome based identification of mouse cumulus cell markers that predict the developmental competence of their enclosed antral oocytes

BACKGROUND: The cumulus cells (CCs) enveloping antral and ovulated oocytes have been regarded as putative source of non-invasive markers of the oocyte developmental competence. A number of studies have indeed observed a correlation between CCs gene expression, embryo quality, and final pregnancy outcome. Here, we isolated CCs from antral mouse oocytes of known developmental incompetence (NSN-CCs) or competence (SN-CCs) and compared their transcriptomes with the aim of identifying distinct marker transcripts. RESULTS: Global gene expression analysis highlighted that both types of CCs share similar transcriptomes, with the exception of 422 genes, 97.6% of which were down-regulated in NSN-CCs vs. SN-CCs. This transcriptional down-regulation in NSN-CCs was confirmed by qRT-PCR analysis of CC-related genes (Has2, Ptx3, Tnfaip6 and Ptgs2). Only ten of the 422 genes were up-regulated with Amh being the most up-regulated in NSN-CCs, with an average 4-fold higher expression when analysed by qRT-PCR. CONCLUSIONS: The developmental incompetence (NSN) or competence (SN) of antral oocytes can be predicted using transcript markers expressed by their surrounding CCs (i.e., Has2, Ptx3, Tnfaip6, Ptgs2 and Amh). Overall, the regulated nature of the group of genes brought out by whole transcriptome analysis constitutes the molecular signature of CCs associated either with developmentally incompetent or competent oocytes and may represent a valuable resource for developing new molecular tools for the assessment of oocyte quality and to further investigate the complex bi-directional interaction occurring between CCs and oocyte

Outcomes of COVID-19 patients treated with continuous positive airway pressure outside ICU

Aim We aim at characterizing a large population of Coronavirus 19 (COVID-19) patients with moderate-to-severe hypoxemic acute respiratory failure (ARF) receiving CPAP outside intensive care unit (ICU), and ascertaining whether the duration of CPAP application increased the risk of mortality for patients requiring intubation. Methods In this retrospective, multicentre cohort study, we included COVID-19 adult patients, treated with CPAP outside ICU for hypoxemic ARF from March 1 st to April 15th, 2020. We collected demographic and clinical data, including CPAP therapeutic goal, hospital length of stay (LOS), and 60- day in-hospital mortality. Results The study includes 537 patients with a median age of 69 (IQR, 60-76) years. Males were 391 (73%). According to predefined CPAP therapeutic goal, 397 (74%) patients were included in full treatment subgroup, and 140 (26%) in the do-not intubate (DNI) subgroup. Median CPAP duration was 4 (IQR, 1-8) days, while hospital LOS 16 (IQR, 9-27) days. Sixty-day in-hospital mortality was overall 34% (95%CI, 0.304-0.384), and 21% (95%CI, 0.169-0.249) and 73% (95%CI, 0.648-0.787) for full treatment and DNI subgroups, respectively. In the full treatment subgroup, in-hospital mortality was 42% (95%CI, 0.345-0.488) for 180 (45%) CPAP failures requiring intubation, while 2% (95%CI, 0.008- 0.035) for the remaining 217 (55%) patients who succeeded. Delaying intubation was associated with increased mortality [HR, 1.093 (95%CI, 1.010-1.184)]. Conclusions We described a large population of COVID-19 patients treated with CPAP outside ICU. Intubation delay represents a risk factor for mortality. Further investigation is needed for early identification of CPAP failures

The role of immune suppression in COVID-19 hospitalization: clinical and epidemiological trends over three years of SARS-CoV-2 epidemic

Specific immune suppression types have been associated with a greater risk of severe COVID-19 disease and death. We analyzed data from patients >17 years that were hospitalized for COVID-19 at the “Fondazione IRCCS Ca′ Granda Ospedale Maggiore Policlinico” in Milan (Lombardy, Northern Italy). The study included 1727 SARS-CoV-2-positive patients (1,131 males, median age of 65 years) hospitalized between February 2020 and November 2022. Of these, 321 (18.6%, CI: 16.8–20.4%) had at least one condition defining immune suppression. Immune suppressed subjects were more likely to have other co-morbidities (80.4% vs. 69.8%, p < 0.001) and be vaccinated (37% vs. 12.7%, p < 0.001). We evaluated the contribution of immune suppression to hospitalization during the various stages of the epidemic and investigated whether immune suppression contributed to severe outcomes and death, also considering the vaccination status of the patients. The proportion of immune suppressed patients among all hospitalizations (initially stable at <20%) started to increase around December 2021, and remained high (30–50%). This change coincided with an increase in the proportions of older patients and patients with co-morbidities and with a decrease in the proportion of patients with severe outcomes. Vaccinated patients showed a lower proportion of severe outcomes; among non-vaccinated patients, severe outcomes were more common in immune suppressed individuals. Immune suppression was a significant predictor of severe outcomes, after adjusting for age, sex, co-morbidities, period of hospitalization, and vaccination status (OR: 1.64; 95% CI: 1.23–2.19), while vaccination was a protective factor (OR: 0.31; 95% IC: 0.20–0.47). However, after November 2021, differences in disease outcomes between vaccinated and non-vaccinated groups (for both immune suppressed and immune competent subjects) disappeared. Since December 2021, the spread of the less virulent Omicron variant and an overall higher level of induced and/or natural immunity likely contributed to the observed shift in hospitalized patient characteristics. Nonetheless, vaccination against SARS-CoV-2, likely in combination with naturally acquired immunity, effectively reduced severe outcomes in both immune competent (73.9% vs. 48.2%, p < 0.001) and immune suppressed (66.4% vs. 35.2%, p < 0.001) patients, confirming previous observations about the value of the vaccine in preventing serious disease

Amh transcript: a non-invasive cumulus cells marker of the oocyte developmental competence

In the field of assisted reproductive technologies (ARTs) there is a growing need to identify non-invasive markers of the oocyte quality. To this end, a number of studies have screened various molecules and morphological characteristics, but reached contrasting and yet not widely accepted results. This study aims at identifying cumulus cell (CC) markers of the oocyte development competence. To this purpose, mouse fully-grown antral follicles were isolated and the CCs separated from the oocytes enclosed. Then, based on their chromatin organization, oocytes were classified as surrounded nucleolus (SN, with a ring of Hoechst-positive heterochromatin surrounding the nucleolus) and non-surrounded nucleolus (NSN, when they displayed spots of heterochromatin widespread within the nucleus). SN oocytes, when matured in vitro to the MII stage, fertilized and the preimplantation embryo transferred in the uterus of a pseudo-pregnant female may reach full term; on the contrary, NSN oocytes arrest development at the 2-cell stage. CCs belonging to SN (SN-CCs) or NSN (NSN-CCs) oocytes were collected separately and analysed. We first made a microarray analysis of the whole transcriptome profile that brought up a list of 422 differentially regulated genes, most of which (97.6%) were down-regulated in NSN-CCs surrounding developmentally incompetent oocytes. A bioinformatics analysis associated these genes to 11 major biological processes, including development and reproduction. In addition, a bibliographic analysis of the literature based on a list of MeSH terms brought out a group of 26 differentially regulated genes involved in ovarian functions. Then, by Real Time PCR (qRT-PCR), we studied the expression of a group of CC-related genes: Has2, Ptx3, Ptgs2 and Tnfaip6, involved in cumulus expansion prior to ovulation and Amh involved in primordial follicles recruitment and dominant follicles selection. In the comparison between NSN-CCs vs. SN-CCs, Has2, Ptx3, Ptgs2 and Tnfaip6 resulted down regulated, whereas Amh showed a strong 4-fold up-regulation, confirming the microarray data. Among the gene sequences highlighted in our study, Amh was clearly the most differentially regulated in the comparison between NSN-CCs vs. SN-CCs, a difference that was confirmed also at the protein level (immunofluorescence). In conclusion, with the use of a model study that allows the identification, a priori, of the developmental capacity of antral mouse oocytes, for the first time, we identified a number of transcripts that could be used as markers of the oocyte developmental competence

Biallelic Inactivation of the Dual Oxidase Maturation Factor 2 (DUOXA2) Gene as a Novel Cause of Congenital Hypothyroidism

Context: Dual oxidase 2 (DUOX2) is the catalytic core of the H2O2 generator crucial for the iodination of thyroglobulin in thyroid hormone synthesis. DUOX2 deficiency produces congenital hypothyroidism (CH) in humans and mice. We recently cloned a novel gene, the product of which (dual oxidase maturation factor 2; DUOXA2) is required to express DUOX2 enzymatic activity