Trefoil factor 2 expressed by the murine pancreatic acinar cells is required for the development of islets and for beta cell function during aging.

Exocrine-to-endocrine crosstalk in the pancreas is crucial to maintain beta cell function. However, the molecular mechanisms underlying this crosstalk are largely undefined. Trefoil factor 2 (Tff2) is a secreted factor known to promote the proliferation of beta cells in vitro, but its physiological role in vivo in the pancreas is unknown. Also, it remains unclear which pancreatic cell type expresses Tff2 protein. We therefore created a mouse model with a conditional knockout of Tff2 in the murine pancreas. We find that the Tff2 protein is preferentially expressed in acinar but not ductal or endocrine cells. Tff2 deficiency in the pancreas reduces beta cell mass on embryonic day 16.5. However, homozygous mutant mice are born without a reduction of beta cells and with acinar Tff3 compensation by day 7. When mice are aged to 1 year, both male and female homozygous and male heterozygous mutants develop impaired glucose tolerance without affected insulin sensitivity. Perifusion analysis reveals that the second phase of glucose-stimulated insulin secretion from islets is reduced in aged homozygous mutant compared to controls. Collectively, these results demonstrate a previously unknown role of Tff2 as an exocrine acinar cell-derived protein required for maintaining functional endocrine beta cells in mice.

Exome sequencing in genuine empty follicle syndrome: Novel candidate genes.

OBJECTIVE(S): Empty follicle syndrome (EFS) is a condition in which no oocytes are retrieved in an IVF cycle despite apparently normal follicular development and meticulous follicular aspiration following ovulation induction. The EFS is called genuine (gEFS) when the trigger administration is correct. The existence of gEFS is a subject of controversy, and it is quite rare with an undetermined etiology. Genetic defects in specific genes have been demonstrated to be responsible for this condition in some patients. Our objective was to identify novel genetic variants associated with gEFS. STUDY DESIGN: We conducted a prospective observational study including 1,689 egg donors from July 2017 to February 2023. WES were performed in patients suffering gEFS. RESULTS: Only 7 patients (0.41 %) exhibited gEFS after two ovarian stimulation cycles and we subsequently performed whole exome sequencing (WES) on these patients. Following stringent filtering, we identified 6 variants in 5 affected patients as pathogenic in new candidate genes which have not been previously associated with gEFS before, but which are involved in important biological processes related to folliculogenesis. These genetic variants included c.603_618del in HMMR, c.1025_1028del in LMNB1, c.1091-1G > A in TDG, c.607C > T in HABP2, c.100 + 2 T > C in HAPLN1 and c.3592_3593del in JAG2. CONCLUSION: As a conclusion, we identified new candidate genes related to gEFS that expand the mutational spectrum of genes related to gEFS.This study show that WES might be an efficient tool to identify the genetic etiology of gEFS and provide further understanding of the pathogenic mechanism of gEFS.

Chemical sensing of common microorganisms found in biopharmaceutical industries using MIR laser spectroscopy and multivariate analysis.

Mid-infrared laser spectroscopy was used to investigate common bacteria encountered in biopharmaceutical industries. The study involved the detection of bacteria using quantum cascade laser spectroscopy coupled to a grazing angle probe (QCL-GAP). Substrates similar to surfaces commonly used in biopharmaceutical industries were used as support media for the samples. Reflectance measurements were assisted by Multivariate Analysis (MVA) to assemble a powerful spectroscopic technique with classification and identification resources. The species analyzed, Staphylococcus aureus, Staphylococcus epidermidis, and Micrococcus luteus, were used to challenge the technique's capability to discriminate from microorganisms of the same family. Principal Components Analysis and Partial Least Squares-Discriminant Analysis differentiated between the bacterial species, using QCL-GAP-MVA as the reference. Spectral differences in the bacterial membrane were used to determine if these microorganisms were present in the samples analyzed. Results herein provided effective discrimination for the bacteria under study with high sensitivity and specificity.

Activation of ductal progenitor-like cells from adult human pancreas requires extracellular matrix protein signaling.

Ductal progenitor-like cells are a sub-population of ductal cells in the adult human pancreas that have the potential to contribute to regenerative medicine. However, the microenvironmental cues that regulate their activation are poorly understood. Here, we establish a 3-dimensional suspension culture system containing six defined soluble factors in which primary human ductal progenitor-like and ductal non-progenitor cells survive but do not proliferate. Expansion and polarization occur when suspension cells are provided with a low concentration (5% v/v) of Matrigel, a sarcoma cell product enriched in many extracellular matrix (ECM) proteins. Screening of ECM proteins identified that collagen IV can partially recapitulate the effects of Matrigel. Inhibition of integrin α1ß1, a major collagen IV receptor, negates collagen IV- and Matrigel-stimulated effects. These results demonstrate that collagen IV is a key ECM protein that stimulates the expansion and polarization of human ductal progenitor-like and ductal non-progenitor cells via integrin α1ß1 receptor signaling.

Impact of Group vs Individual Embryo Culture Strategies on Blastocyst and Clinical Outcomes.

Embryo culture is one of the most important steps in an assisted reproduction laboratory. Embryos can be cultured individually, one embryo per media drop, or in groups, culturing several embryos in the same media drop. Due to the controversy generated on this subject, we wondered which embryo culture method would have the best results in terms of quality and blastocyst formation rate. We designed a prospective randomized study comparing two different embryo culture strategies: group and individual embryo culture. The data were obtained from 830 embryos from 103 egg donation treatments. The zygotes were randomized into two groups: individual culture (group 1) or group culture (group 2). The embryos were cultured in 35-µl drops until day 5 when they were classified morphologically. We observed a significant increase in the blastocyst formation rate and in the usable embryo rate in individual culture on day 5 compared to group culture. However, good embryo quality (A/B blastocysts), implantation, and pregnancy rates were similar regardless of the type of embryo-culture. As a conclusion, individual culture may increase blastocyst formation rate and may benefit embryo quality on day 5. Our results support previous reports suggesting that individual culture could improve embryo development.

Rare, Tightly-Bound, Multi-Cellular Clusters in the Pancreatic Ducts of Adult Mice Function Like Progenitor Cells and Survive and Proliferate After Acinar Cell Injury.

Pancreatic ductal progenitor cells have been proposed to contribute to adult tissue maintenance and regeneration after injury, but the identity of such ductal cells remains elusive. Here, from adult mice, we identify a near homogenous population of ductal progenitor-like clusters, with an average of 8 cells per cluster. They are a rare subpopulation, about 0.1% of the total pancreatic cells, and can be sorted using a fluorescence-activated cell sorter with the CD133highCD71lowFSCmid-high phenotype. They exhibit properties in self-renewal and tri-lineage differentiation (including endocrine-like cells) in a unique 3-dimensional colony assay system. An in vitro lineage tracing experiment, using a novel HprtDsRed/+ mouse model, demonstrates that a single cell from a cluster clonally gives rise to a colony. Droplet RNAseq analysis demonstrates that these ductal clusters express embryonic multipotent progenitor cell markers Sox9, Pdx1, and Nkx6-1, and genes involved in actin cytoskeleton regulation, inflammation responses, organ development, and cancer. Surprisingly, these ductal clusters resist prolonged trypsin digestion in vitro, preferentially survive in vivo after a severe acinar cell injury and become proliferative within 14 days post-injury. Thus, the ductal clusters are the fundamental units of progenitor-like cells in the adult murine pancreas with implications in diabetes treatment and tumorigenicity.

Conventional ovarian stimulation vs. delayed single dose corifollitropin alfa ovarian stimulation in oocyte donors: a prospective randomized study. Tail trial.

The present study compares two protocols for ovarian controlled stimulation in terms of number of cumulus-oocyte complexes and metaphase II oocytes. We employed a single injection of 150mcg of corifollitropin alfa after a 7-day oral contraceptive pill-free interval for TAIL group and a conventional administration of corifollitropin alfa after a 5-day OCP-free interval with additional rFSH from 8th of ovarian controlled stimulation. Prospective, randomized, comparative, non-inferiority, opened and controlled trial carried out in 180 oocyte donors 31 were excluded, 81 were randomized to the control group and 68 to the TAIL group. No differences were found in the number of follicles larger than 14 and 17 mm at triggering day. However, a lower number of cumulus-oocyte complexes and metaphase II oocytes were obtained in TAIL group compared to the control group, expressed as median (interquartile range): 10.5 (5.5-19) vs. 14 [11-21] and 9 (4-13) vs. 12 (9-17) respectively. Additionally, the incidence of failed retrieval or metaphase II oocytes = 0 was higher in TAIL group 7(10.3%) vs. 1(1.2%) p = 0.024. The use of a single injection of corifollitropin alfa after a 7-day oral contraceptive pill-free interval in oocyte donors resulted in a lower number of cumulus-oocyte complexes and metaphase II oocytes. No additional rFSH was administered in this group. Clinical Trial Registration: https://www.clinicaltrialsregister.eu/ctr-search/trial/2019-001343-44/results.

Influence of the starting day of luteal phase stimulation on double stimulation cycles.

Background: Double ovarian stimulation is one of the most used strategies in poor-prognosis patients. There is a high heterogeneity between the studies regarding the execution of this stimulation protocol. The aim of this study was to investigate whether the day on which luteal phase stimulation begins after the first oocyte retrieval affects ovarian response in DuoStim cycles. Methods: This observational and retrospective study included 541 DuoStim cycles between January 2018 and December 2021 in a private fertility clinic. Patients were assigned to 4 groups according to the timing of the onset of luteal phase stimulation after oocyte retrieval (0-2nd day, 3rd day, 4th day and 5th-6th day). The primary outcome was the number of oocytes retrieved in the luteal phase in each group. Results: No differences were found between groups in the number of oocytes collected (5.12 ± 3.56 vs. 5.39 ± 3.74 vs. 5.61 ± 3.94 vs. 5.89 ± 3.92; p=0,6), MII or number of follicles. An increase in the duration of stimulation was found when stimulation started on the 4th day (10.42 ± 2.31 vs. 10.68 ± 2.37 vs. 11.27 ± 2.40 vs. 10.65 ± 2.37 days, p=0,033). A lower number of fertilized oocytes was observed when stimulation began before the fourth day (3.36 ± 2.80 vs. 3.95 ± 2.53 vs. 4.03 ± 2.73 vs. 4.48 ± 3.11; p=0,036). The number of blastocysts was higher when the stimulation started 5-6 days after retrieval (1.82 ± 1.74 vs. 2.13 ± 1.61 vs. 2.33 ± 2.06 vs. 2.91 ± 2.39; p= 0,030). Discussion: The number of oocytes retrieved does not differ depending on the day that stimulation begins. However, oocytes competence in terms of fertilized oocytes and blastulation, appears to be lower when the second stimulation starts before the fourth day after oocyte retrieval.

Methylcellulose colony assay and single-cell micro-manipulation reveal progenitor-like cells in adult human pancreatic ducts.

Progenitor cells capable of self-renewal and differentiation in the adult human pancreas are an under-explored resource for regenerative medicine. Using micro-manipulation and three-dimensional colony assays we identify cells within the adult human exocrine pancreas that resemble progenitor cells. Exocrine tissues were dissociated into single cells and plated into a colony assay containing methylcellulose and 5% Matrigel. A subpopulation of ductal cells formed colonies containing differentiated ductal, acinar, and endocrine lineage cells, and expanded up to 300-fold with a ROCK inhibitor. When transplanted into diabetic mice, colonies pre-treated with a NOTCH inhibitor gave rise to insulin-expressing cells. Both colonies and primary human ducts contained cells that simultaneously express progenitor transcription factors SOX9, NKX6.1, and PDX1. In addition, in silico analysis identified progenitor-like cells within ductal clusters in a single-cell RNA sequencing dataset. Therefore, progenitor-like cells capable of self-renewal and tri-lineage differentiation either pre-exist in the adult human exocrine pancreas, or readily adapt in culture.

Exploring self-detection of the endogenous LH surge using a urine test as a tool to predict a suboptimal response to gonadotropin-releasing hormone agonist trigger during in vitro fertilization cycles.

OBJECTIVE: Is self-detection of the endogenous LH surge using a urine testing a reliable method to confirm a successful gonadotropin-releasing hormone agonist (GnRHa) trigger in IVF cycles? METHODS: Prospective observational study including a total of 103 oocyte donation cycles between November 2019 and January 2020. Urine LH testing (Akralab SL, Spain, cut-of value 30 mIU/mL) was performed at home in samples from the first micturition in the morning after the GnRHa trigger and a picture of the result was sent to the nurse coordinator; this information was concealed and only disclosed after oocyte aspiration. RESULTS: From the total group, two cycles were excluded. A total of 101 oocyte donors performed the LH urine testing, all proceeded to oocyte aspiration and were included in final analysis. A total of 85 (84.2%) had a positive LH test and an uneventful oocyte retrieval with good retrieval rates (false positive rate: 0%). A total of 16 had a negative LH test (15.8%) and had a good oocyte retrieval rates (false negative rate: 15.8%). There were no cases of empty follicle syndrome. CONCLUSIONS: Due to a high false negative rate, self-testing of endogenous LH release using a LH urine test when performed approximately 12-hours after triggering does not seem to be a reliable method to predict a suboptimal response to gonadotropin-releasing hormone.

Colored sticky traps for monitoring phytophagous thrips (Thysanoptera) in mango agroecosystems, and their impact on beneficial insects.

The capture efficiency of six colored sticky traps (blue, green, orange, purple, white, and yellow) was tested in mango agroecosystems of Mexico with the purpose to: (i) document the diversity of Thysanoptera; (ii) determine the attraction of phytophagous thrips; (iii) assess the impact of these traps on beneficial insects; and (iv) assess the relationship between the density of Frankliniella thrips captured on traps and those found in the inflorescences. The use of colored sticky traps has revealed a great diversity of thrips and beneficial insects in the mango agroecosystem. A total of 16,441 thrips were caught on sticky traps throughout the sampling period, of which 16,251 (98.8%) were thrips adults and 190 (1.2%) larvae. Forty one species of thrips were collected either from sticky traps or from inflorescences. Of these, 32 species feed either on leaves or flowers. Frankliniella cephalica, F. gardeniae and F. invasor, were the most abundant species. Scirtothrips citri and S. manihoti were also captured among other phytophagous thrips. The white trap captured significantly more Frankliniella species and also had the smallest capture of beneficial insects. Yellow traps were the most attractive for Scirtothrips species, with low detrimental effects on insect pollinators, although high impact on natural enemies. Thrips species captured on sticky traps showed a low and non-significantly correlation with respect to the density of thrips in mango inflorescences. Although sticky traps did not predict the density of Frankliniella populations in mango inflorescences, the study represents a substantial progress in the use of color traps in mango agroecosystems. Colored sticky traps would be a good option for monitoring mango thrips to detect them at earlier stages of infestation to implement management tactics and avoid the building-up of thrips populations.

Application of machine learning to predict aneuploidy and mosaicism in embryos from in vitro fertilization cycles.

BACKGROUND: The factors associated with embryo aneuploidy have been extensively studied. Mostly maternal age and to a lesser extent male factor and ovarian stimulation have been related to the occurrence of chromosomal alterations in the embryo. On the other hand, the main factors that may increase the incidence of embryo mosaicism have not yet been established. OBJECTIVE: This study aimed to establish a machine learning model that would allow prediction of aneuploidies and mosaicism in embryos conceived via in vitro fertilization, and thus help to determine which variables are associated with these chromosomal alterations. STUDY DESIGN: The study design was observational and retrospective. A total of 6989 embryos from 2476 cycles of preimplantation genetic testing for aneuploidies were included (January 2013 to December 2020). The trophoectoderm biopsies on day-5, -6, or -7 blastocysts were analyzed by preimplantation genetic testing for aneuploidies (PGT-A). The different maternal, paternal, couple, embryo, and in vitro fertilization cycle characteristics were recorded in a database (22 predictor variables) from which predictive models of embryo aneuploidy and mosaicism were developed; 16 different unsupervised classification machine learning algorithms were used to establish the predictive models. RESULTS: Two different predictive models were performed: one for aneuploidy and the other for mosaicism. The predictor variable was of multiclass type because it included the segmental- and whole-chromosome alteration categories. The best predicting models for both aneuploidies and mosaicism were those obtained from the Random Forest algorithm. The area under ROC curve (AUC) value was 0.792 for the aneuploidy explanatory model and 0.776 for mosaicism. The most important variable in the final aneuploidy model was maternal age, followed by paternal and maternal karyotype and embryo quality. In the predictive model of mosaicism, the most important variable was the technique used in preimplantation genetic testing for aneuploidies and embryo quality, followed by maternal age and day of biopsy. CONCLUSION: It is possible to predict embryo aneuploidy and mosaicism from certain characteristics of the patients and their embryos.

Computer Numerical Control Micromilling of a Microfluidic Acrylic Device with a Staggered Restriction for Magnetic Nanoparticle-based Immunoassays.

Microfluidic systems have greatly improved immunoassay techniques. However, many microfabrication techniques require specialized, expensive, or complicated equipment, making fabrication costly and incompatible with mass production, which is one of the most important preconditions for point-of-care tests (POCT) to be adopted in low-resource settings. This work describes the fabrication process of an acrylic (polymethylmethacrylate, PMMA) device for nanoparticle-conjugated enzymatic immunoassay testing using the computer numerical control (CNC) micromilling technique. The functioning of the microfluidic device is shown by performing an immunoassay to detect a commercial antibody using lysozyme as a model antigen conjugated to 100 nm magnetic nanoparticles. This device integrates a physical staggered restriction of only 5 µm in height, used to capture magnetic microparticles that make up a magnetic trap by placing an external magnet. In this way, the magnetic force on the immunosupport of conjugated nanoparticles is enough to capture them and resist flow drag. This microfluidic device is particularly suitable for low-cost mass production without the loss of precision for immunoassay performance.

Approach to the Fatigue and Cellular Behavior of Superficially Modified Porous Titanium Dental Implants.

In this work, the fatigue and cellular performance of novel superficially treated porous titanium dental implants made up using conventional powder metallurgy and space-holder techniques (30 vol.% and 50 vol.%, both with a spacer size range of 100-200 µm) are evaluated. Before the sintering stage, a specific stage of CNC milling of the screw thread of the implant is used. After the consolidation processing, different surface modifications are performed: chemical etching and bioactive coatings (BG 45S5 and BG 1393). The results are discussed in terms of the effect of the porosity, as well as the surface roughness, chemical composition, and adherence of the coatings on the fatigue resistance and the osteoblast cells' behavior for the proposed implants. Macro-pores are preferential sites of the nucleation of cracks and bone cell adhesion, and they increase the cellular activity of the implants, but decrease the fatigue life. In conclusion, SH 30 vol.% dental implant chemical etching presents the best bio-functional (in vitro osseointegration) and bio-mechanical (stiffness, yield strength and fatigue life) balance, which could ensure the required characteristics of cortical bone tissue.

Using Nonword Repetition to Identify Language Impairment in Bilingual Children: A Meta-Analysis of Diagnostic Accuracy.

Purpose Nonword repetition has been endorsed as a less biased method of assessment for children from culturally and linguistically diverse backgrounds, but there are currently no systematic reviews or meta-analyses on its use with bilingual children. The purpose of this study was to evaluate diagnostic accuracy of nonword repetition in the identification of language impairment (LI) in bilingual children. Method Using a key word search of peer-reviewed literature from several large electronic databases, as well as ancestral and forward searches, 13 studies were identified that met the eligibility criteria. Studies were evaluated on the basis of quality of evidence, design characteristics, and reported diagnostic accuracy. A meta-regression analysis, based on study results, was conducted to identify task characteristics that may be associated with better classification accuracy. Results Diagnostic accuracy across studies ranged from poor to good. Bilingual children with LI performed with more difficulty on nonword repetition tasks than those with typical language. Quasi-universal tasks, which account for the phonotactic constraints of multiple languages, exhibited better diagnostic accuracy and resulted in less misidentification of children with typical language than language-specific tasks. Conclusions Evidence suggests that nonword repetition may be a useful tool in the assessment and screening of LI in bilingual children, though it should be used in conjunction with other measures. Quasi-universal tasks demonstrate the potential to further reduce assessment bias, but extant research is limited.

Nursing Care at the Service of Life: A model including charity, quality, and leadership.

BACKGROUND: Nursing professional practice models allow the focus on care and qualification of hospital service by articulating the theory to the practice of nursing care with the final purpose of improving the patient's and family caregivers' quality of life. OBJECTIVE: To describe the model "Nursing care at the service of life," built and validated in a Colombian clinic, 2019. METHOD: A Nursing Methodology Research developed in four consecutive steps intended to identify the need to have a theoretical framework; to build it and validate it in a collective way that included the different members of the clinic nursing staff. RESULTS: The main concepts and articulated assumptions of the nursing practice model were described. The model will help the nursing staff to achieve the institutional mission. CONCLUSIONS: The professional practice Model "Nursing care at the service of life" reflects the nursing profession's charity, quality, and leadership, as a hospital care axis. The institution's nursing staff were able to identify themselves in the model's approach, also the expert criteria endorsed the construction process and content. The validation process done with the nurse's staff demonstrates that this model is accepted and useful to guide the nursing caring practices within the clinic.

Methylenetetrahydrofolate reductase gene polymorphisms are not associated with embryo chromosomal abnormalities and IVF outcomes.

The aim of our study was to investigate the effect of maternal and embryo MTHFR C677T and A1298C polymorphisms on embryo aneuploidies and mosaicism and the correlation between these genetic variants in transferred euploid embryos and IVF outcomes. MTHFR genotype was analyzed in 77 women who performed an IVF/ICSI cycle with PGT-A. Moreover, to evaluate the effect of embryo MTHFR polymorphisms on embryo aneuploidies and mosaicism, the MTHFR genotype was analyzed in 191 biopsied embryos from the PGT-A cycles of these patients. Additionally, 218 DNA samples from trophectoderm biopsies belonging to a different group of patients were also genotyped. MTHFR polymorphisms were analyzed in a total amount of 409 trophectoderm samples. The main parameters analyzed were embryo aneuploidy and mosaicism rates. Finally, the IVF outcomes of 241 single euploid embryo transfers were assessed and compared between different MTHFR embryo genotypes. The aneuploidy rates were similar in embryos from homozygous normal women and women with at least one mutated allele (54.7% vs. 30.2% in 677C>T and 37.8% vs. 42.7% in 1298A>C). Furthermore, no differences were observed in the mosaicism rate (24.0% vs. 13.8% in 677C>T and 17.1% vs. 17.3% in 1298A>C). A similar analysis was performed, taking into account the embryo genotype results. No differences in aneuploidy rate were observed between the study groups. The only significant difference was the mosaicism rate among 677C>T genotype (13.5% in 677CC group vs. 5.4% in 677CT/TT; p = 0.019). Implantation rate, biochemical and clinical miscarriage rates, and ongoing pregnancy rate were compared between different embryo genotypes, and no statistically significant differences were found. In conclusion, the maternal MTHFR genotype did not influence embryo chromosomal abnormalities. Moreover, the embryo MTHFR genotype was not associated with embryo aneuploidy or IVF outcomes such as implantation, pregnancy loss, and ongoing pregnancy when euploid embryos were transferred.Abbreviations: MTHFR: methylenetetrahydrofolate reductase; IVF: in vitro fertilization; PGT-A: preimplantation genetic testing for aneuploidies; SAM: S-adenosyl methionine; SNP: single nucleotide polymorphism; SPSS: Statistical Package for Social Sciences; RIF: recurrent implantation failure; RPL: recurrent pregnancy loss; hCG: human chorionic gonadotropin; PBS: phosphate buffered saline; CGH: comparative genomic hybridization; NGS: next generation sequencing.