Synthetic Par polarity induces cytoskeleton asymmetry in unpolarized mammalian cells.

Polarized cells rely on a polarized cytoskeleton to function. Yet, how cortical polarity cues induce cytoskeleton polarization remains elusive. Here, we capitalized on recently established designed 2D protein arrays to ectopically engineer cortical polarity of virtually any protein of interest during mitosis in various cell types. This enables direct manipulation of polarity signaling and the identification of the cortical cues sufficient for cytoskeleton polarization. Using this assay, we dissected the logic of the Par complex pathway, a key regulator of cytoskeleton polarity during asymmetric cell division. We show that cortical clustering of any Par complex subunit is sufficient to trigger complex assembly and that the primary kinetic barrier to complex assembly is the relief of Par6 autoinhibition. Further, we found that inducing cortical Par complex polarity induces two hallmarks of asymmetric cell division in unpolarized mammalian cells: spindle orientation, occurring via Par3, and central spindle asymmetry, depending on aPKC activity.

Macromolecular condensation buffers intracellular water potential.

Optimum protein function and biochemical activity critically depends on water availability because solvent thermodynamics drive protein folding and macromolecular interactions1. Reciprocally, macromolecules restrict the movement of 'structured' water molecules within their hydration layers, reducing the available 'free' bulk solvent and therefore the total thermodynamic potential energy of water, or water potential. Here, within concentrated macromolecular solutions such as the cytosol, we found that modest changes in temperature greatly affect the water potential, and are counteracted by opposing changes in osmotic strength. This duality of temperature and osmotic strength enables simple manipulations of solvent thermodynamics to prevent cell death after extreme cold or heat shock. Physiologically, cells must sustain their activity against fluctuating temperature, pressure and osmotic strength, which impact water availability within seconds. Yet, established mechanisms of water homeostasis act over much slower timescales2,3; we therefore postulated the existence of a rapid compensatory response. We find that this function is performed by water potential-driven changes in macromolecular assembly, particularly biomolecular condensation of intrinsically disordered proteins. The formation and dissolution of biomolecular condensates liberates and captures free water, respectively, quickly counteracting thermal or osmotic perturbations of water potential, which is consequently robustly buffered in the cytoplasm. Our results indicate that biomolecular condensation constitutes an intrinsic biophysical feedback response that rapidly compensates for intracellular osmotic and thermal fluctuations. We suggest that preserving water availability within the concentrated cytosol is an overlooked evolutionary driver of protein (dis)order and function.

Oxidized cholesteryl ester induces exocytosis of dysfunctional lysosomes in lipidotic macrophages.

A key event in atherogenesis is the formation of lipid-loaded macrophages, lipidotic cells, which exhibit irreversible accumulation of undigested modified low-density lipoproteins (LDL) in lysosomes. This event culminates in the loss of cell homeostasis, inflammation, and cell death. Nevertheless, the exact chemical etiology of atherogenesis and the molecular and cellular mechanisms responsible for the impairment of lysosome function in plaque macrophages are still unknown. Here, we demonstrate that macrophages exposed to cholesteryl hemiazelate (ChA), one of the most prevalent products of LDL-derived cholesteryl ester oxidation, exhibit enlarged peripheral dysfunctional lysosomes full of undigested ChA and neutral lipids. Both lysosome area and accumulation of neutral lipids are partially irreversible. Interestingly, the dysfunctional peripheral lysosomes are more prone to fuse with the plasma membrane, secreting their undigested luminal content into the extracellular milieu with potential consequences for the pathology. We further demonstrate that this phenotype is mechanistically linked to the nuclear translocation of the MiT/TFE family of transcription factors. The induction of lysosome biogenesis by ChA appears to partially protect macrophages from lipid-induced cytotoxicity. In sum, our data show that ChA is involved in the etiology of lysosome dysfunction and promotes the exocytosis of these organelles. This latter event is a new mechanism that may be important in the pathogenesis of atherosclerosis.

Cost-effectiveness of active tuberculosis screening among high-risk populations in low tuberculosis incidence countries: a systematic review, 2008 to 2023.

BackgroundIn countries with a low TB incidence (≤ 10 cases/100,000 population), active pulmonary tuberculosis (PTB) mostly affects vulnerable populations with limited access to healthcare. Thus, passive case-finding systems may not be successful in detecting and treating cases and preventing further transmission. Active and cost-effective search strategies can overcome this problem.AimWe aimed to review the evidence on the cost-effectiveness (C-E) of active PTB screening programmes among high-risk populations in low TB incidence countries.MethodsWe performed a systematic literature search covering 2008-2023 on PubMed, Embase, Center for Reviews and Dissemination, including Database of Abstracts of Reviews of Effects (DARE), National Health Services Economic Evaluation Database (NHS EED), Global Index Medicus and Cochrane Central Register of Controlled Trials (CENTRAL).ResultsWe retrieved 6,318 articles and included nine in this review. All included studies had an active case-finding approach and used chest X-ray, tuberculin skin test, interferon-gamma release assay and a symptoms questionnaire for screening. The results indicate that screening immigrants from countries with a TB incidence > 40 cases per 100,000 population and other vulnerable populations as individuals from isolated communities, people experiencing homelessness, those accessing drug treatment services and contacts, is cost-effective in low-incidence countries.ConclusionIn low-incidence countries, targeting high-risk groups is C-E. However, due to the data heterogenicity, we were unable to compare C-E. Harmonisation of the methods for C-E analysis is needed and would facilitate comparisons. To outline comprehensive screening and its subsequent C-E analysis, researchers should consider multiple factors influencing screening methods and outcomes.

Anatomical outcomes and complications of sacrocolpopexy using Surelift Uplift mesh: A multicentric observational study.

OBJECTIVE: The study evaluated the anatomical and functional outcomes, as well as the safety data of laparoscopic sacrocolpopexy (LSC) for pelvic organ prolapse (POP) using a lightweight macroporous mesh. METHODS: A multicentric observational study was developed including five expert centers between March 2011 and December 2019. Inclusion criteria were female patients with symptomatic ≥stage II POP (POP-Q classification), who underwent a LSC. A lightweight and macroporous mesh device (Surelift Uplift) was used. Baseline anatomical positions were evaluated using POP-Q stage. The anatomical outcomes and procedural complications were assessed during the postoperative period. Primary outcomes were anatomical success, defined as POP-Q stage ≤I, and subjective success, defined as no bothersome bulge symptoms, and no repeat surgery or pessary use for recurrent prolapse. RESULTS: A total of 325 LSCs were analyzed with a median patient age of 66 (interquartile range [IQR] 61-73). After a median follow-up of 68 months (IQR 46.5-89), anatomical success was found in 88.9%, whereas subjective success was seen in 98.5% of the patients. Recurrent prolapse presented as cystocele (1.5%). Reported complications were bladder (4.6%) or rectum lesions (0.6%), de novo urinary incontinence (12.9%), and mesh extrusion (1.2%). CONCLUSIONS: LSC provides significant clinical improvement and excellent anatomical results, with a low risk of serious complications for women with ≥2 grade POP in a real clinical practice setting.

TATA-Binding Protein-Based Virtual Screening of FDA Drugs Identified New Anti-Giardiasis Agents.

Parasitic diseases, predominantly prevalent in developing countries, are increasingly spreading to high-income nations due to shifting migration patterns. The World Health Organization (WHO) estimates approximately 300 million annual cases of giardiasis. The emergence of drug resistance and associated side effects necessitates urgent research to address this growing health concern. In this study, we evaluated over eleven thousand pharmacological compounds sourced from the FDA database to assess their impact on the TATA-binding protein (TBP) of the early diverging protist Giardia lamblia, which holds medical significance. We identified a selection of potential pharmacological compounds for combating this parasitic disease through in silico analysis, employing molecular modeling techniques such as homology modeling, molecular docking, and molecular dynamics simulations. Notably, our findings highlight compounds DB07352 and DB08399 as promising candidates for inhibiting the TBP of Giardia lamblia. Also, these compounds and DB15584 demonstrated high efficacy against trophozoites in vitro. In summary, this study identifies compounds with the potential to combat giardiasis, offering the prospect of specific therapies and providing a robust foundation for future research.

The structure of root-associated fungal communities is related to the long-term effects of plant diversity on productivity.

Root-associated fungi could play a role in determining both the positive relationship between plant diversity and productivity in experimental grasslands, and its strengthening over time. This hypothesis assumes that specialized pathogenic and mutualistic fungal communities gradually assemble over time, enhancing plant growth more in species-rich than in species-poor plots. To test this hypothesis, we used high-throughput amplicon sequencing to characterize root-associated fungal communities in experimental grasslands of 1 and 15 years of age with varying levels of plant species richness. Specifically, we tested whether the relationship between fungal communities and plant richness and productivity becomes stronger with the age of the experimental plots. Our results showed that fungal diversity increased with plant diversity, but this relationship weakened rather than strengthened over the two time points. Contrastingly, fungal community composition showed increasing associations with plant diversity over time, suggesting a gradual build-up of specific fungal assemblages. Analyses of different fungal guilds showed that these changes were particularly marked in pathogenic fungi, whose shifts in relative abundance are consistent with the pathogen dilution hypothesis in diverse plant communities. Our results suggest that root-associated fungal pathogens play more specific roles in determining the diversity-productivity relationship than other root-associated plant symbionts.

Comparative anatomy of the ovine and human pelvis for laparoscopic sacrocolpopexy: evaluating the effectiveness of the ovine model.

INTRODUCTION AND HYPOTHESIS: Laparoscopic sacrocolpopexy (LSC) is a functional reconstructive surgery used to treat pelvic organ prolapse (POP) in middle-aged women. Although LSC is widely used, its implementation is hindered by perceived technical difficulties and surgical learning curves. Surgeons require adequate experience with LSC prior to performing the procedure on patients to improve their quality of life. This study is aimed at demonstrating the effectiveness of the ovine model (OM) for training and research in LSC, while also comparing anatomical differences between ovine and human models during the procedure. METHODS: The animal model and training were provided by the Jesús Usón Minimally Invasive Surgery Centre. Urologists and gynecologists with experience in LSC participated in a course and their findings were recorded and documented. RESULTS: Differences in patient positioning, trocar placement, and reperitonealization were identified between the ovine and human models. Hysterectomy is always performed in the ovine model, whereas it is not mandatory in humans. There are also differences in the dissection of the levator ani muscle and attachment point of the posterior mesh to the uterus between the two models. Despite differences in some areas, the ovine pelvic structure and vagina are similar in size to those of humans. CONCLUSIONS: The ovine model is a valuable tool for surgeons in their learning curve for LSC, allowing for safe and effective practice prior to performing the procedure on patients. The use of the OM can help to improve the quality of life for women affected by pelvic organ prolapse.

Predictive value of texture analysis on lumbar MRI in patients with chronic low back pain.

PURPOSE: The aim of this study was to determine whether MRI texture analysis could predict the prognosis of patients with non-specific chronic low back pain. METHODS: A prospective observational study was conducted on 100 patients with non-specific chronic low back pain, who underwent a conventional MRI, followed by rehabilitation treatment, and revisited after 6 months. Sociodemographic variables, numeric pain scale (NPS) value, and the degree of disability as measured by the Roland-Morris disability questionnaire (RMDQ), were collected. The MRI analysis included segmentation of regions of interest (vertebral endplates and intervertebral disks from L3-L4 to L5-S1, paravertebral musculature at the L4-L5 space) to extract texture variables (PyRadiomics software). The classification random forest algorithm was applied to identify individuals who would improve less than 30% in the NPS or would score more than 4 in the RMDQ at the end of the follow-up. Sensitivity, specificity, and the area under the ROC curve were calculated. RESULTS: The final series included 94 patients. The predictive model for classifying patients whose pain did not improve by 30% or more offered a sensitivity of 0.86, specificity 0.57, and area under the ROC curve 0.71. The predictive model for classifying patients with a RMDQ score 4 or more offered a sensitivity of 0.83, specificity of 0.20, and area under the ROC curve of 0.52. CONCLUSION: The texture analysis of lumbar MRI could help identify patients who are more likely to improve their non-specific chronic low back pain through rehabilitation programs, allowing a personalized therapeutic plan to be established.

Factors affecting the in vitro embryo production in buffalo (Bubalus bubalis): A review.

Under natural and well-managed conditions, the buffalo has good reproductive and productive indices. However, in vitro embryo production (IVEP) has been used commercially to maximise the number of elite animals. In this species, several factors (donor management, in vitro culture medium, semen, in vitro conditions, embryo transfer) still affect the IVEP results. In addition, the cost of this technique is very high for this purpose. Therefore, more studies, as well as adequate plans, are needed to achieve this objective efficiently. In this review, we discussed the current commercial status, influencing factors (in vivo and in vitro), and the progress and future challenges of IVEP in buffalo. A total of 81 references were used from 1979 to 2022. The relevant data or literature were searched using the following databases: Google, ResearchGate, Science Alert, Science Direct and PubMed, using the following keywords: buffalo oocytes/COCs, buffalo embryos, pregnancy and calving or live birth rate after embryo transfer. The best maturation, cleavage and blastocyst rates in the in vitro production of buffalo embryos were 95.8, 75.2 and 33.4%, respectively. The pregnancy and live birth rates ranged from 22.2% to 43.5% and from 15.3% to 36.5%, respectively, after the transfer of fresh embryos produced in vitro to the recipients. This review will help to contextualise IVEP in buffaloes, as well as create an adequate plan for implementing IVEP in buffaloes.

Does the addition of follicular fluid in the in vitro maturation medium increase the oocyte maturation and embryo production in alpacas?

In alpacas (Vicugna pacos), the high cost of in vitro embryo production is also a consequence of the use of several substances in the culture medium. In addition, embryo production rates in this species are still considered low. Thus, in attempt to reduce the cost and to improve the in vitro embryo production rates, this study evaluates the effect of adding follicular fluid (FF) in the in vitro maturation medium on oocyte maturation and subsequent embryo production. After ovary collection at the local slaughterhouse, the oocytes were recovered, selected, and allocated in experimental groups: standard maturation medium (G1) and simplified medium added by 10% FF (G2). The FF was acquired from follicles between 7- and 12-mm diameter. The cumulus cell expansion and the embryo production rates were analyzed by chi-square with p < 0.05. No differences (p > 0.05) were observed in maturation rate between G1 (66.36%) and G2 (63.12%) groups. Likewise, no significant difference (p > 0.05) was verified between G1 and G2 for morula (40.85 vs 38.45%), blastocyst (7.01 vs 6.93%), and total number of embryos (47.87 vs 45.38%). In conclusion, it was possible to simplify the medium used for in vitro maturation of alpaca oocytes resulting in embryo production rates similar to the standard medium.

How reactive is investment in US green bonds and ESG-eligible stocks in times of crisis? Exploring the COVID-19 crisis.

This study examines how green bonds and environmental, social and governance (ESG) stock market returns have reacted to the COVID-19 crisis in the US. Unlike the Standard and Poor's (S&P) 500 index, the response of green bonds and ESG markets to pandemic progress is nonlinear: A low (large) level of confirmed new cases of COVID-19 has a positive (negative) impact. Furthermore, the implemented containment policies (stringency measures and vaccination campaigns) are positively valued, but their simultaneous use is perceived by investors as a bad signal. Overall, our findings question the resilience of investments in green bonds and ESG markets.

Deciphering the role of specialist and generalist plant-microbial interactions as drivers of plant-soil feedback.

Feedback between plants and soil microbial communities can be a powerful driver of vegetation dynamics. Plants elicit changes in the soil microbiome that either promote or suppress conspecifics at the same location, thereby regulating population density-dependence and species co-existence. Such effects are often attributed to the accumulation of host-specific antagonistic or beneficial microbiota in the rhizosphere. However, the identity and host-specificity of the microbial taxa involved are rarely empirically assessed. Here we review the evidence for host-specificity in plant-associated microbes and propose that specific plant-soil feedbacks can also be driven by generalists. We outline the potential mechanisms by which generalist microbial pathogens, mutualists and decomposers can generate differential effects on plant hosts and synthesize existing evidence to predict these effects as a function of plant investments into defence, microbial mutualists and dispersal. Importantly, the capacity of generalist microbiota to drive plant-soil feedbacks depends not only on the traits of individual plants but also on the phylogenetic and functional diversity of plant communities. Identifying factors that promote specialization or generalism in plant-microbial interactions and thereby modulate the impact of microbiota on plant performance will advance our understanding of the mechanisms underlying plant-soil feedback and the ways it contributes to plant co-existence.

Efficacy and safety of abciximab versus tirofiban in addition to ticagrelor in STEMI patients undergoing primary percutaneous intervention.

Platelet glycoprotein IIb/IIIa inhibitors (GPIs) have been part of the adjuvant treatment of acute coronary syndrome for years. However, real-life data regarding the efficacy and safety of GPIs under the current indications are lacking in the setting of potent platelet inhibition. The objectives were to assess the efficacy and safety of abciximab versus tirofiban in patients with ST-elevation acute myocardial infarction (STEMI) undergoing primary percutaneous coronary intervention (PPCI) and pretreated with ticagrelor, and to identify independent predictor factors of efficacy, bleeding and platelet drop. Three hundred sixty-two patients were divided by GPI administered. Clinical, laboratory, angiographic and outcome characteristics were compared. The primary objective was a composite efficacy endpoint (death from any cause, nonfatal myocardial infarction and nonfatal stroke) at 30 days. The secondary objectives were its individual components, safety (bleeding) and the impact on platelet count during hospital stay. The composite efficacy endpoint was similar in the abciximab and tirofiban groups (6.1% vs 7.3%; p = .632). There were also no differences in cardiovascular death (2.5% vs 2.4%; p = .958), nonfatal myocardial infarction (3% vs 4.3%; p = .521) and nonfatal stroke (0.5% vs 1.8%; p = .332). Tirofiban administration was associated with a higher incidence of bleeding (11.6% vs 22%; p = .008) with no differences in BARC ≥ 3b bleeding (3.6 vs 2.5%; p = .760). In STEMI patients undergoing PPCI with ticagrelor, abciximab and tirofiban had similar rates in the composite efficacy endpoint at 30 days. The 30-day bleeding rate was significantly higher in the tirofiban group. Tirofiban administration was an independent predictor of both bleeding and platelet count drop.

Combination of Feature Selection and Resampling Methods to Predict Preterm Birth Based on Electrohysterographic Signals from Imbalance Data.

Due to its high sensitivity, electrohysterography (EHG) has emerged as an alternative technique for predicting preterm labor. The main obstacle in designing preterm labor prediction models is the inherent preterm/term imbalance ratio, which can give rise to relatively low performance. Numerous studies obtained promising preterm labor prediction results using the synthetic minority oversampling technique. However, these studies generally overestimate mathematical models' real generalization capacity by generating synthetic data before splitting the dataset, leaking information between the training and testing partitions and thus reducing the complexity of the classification task. In this work, we analyzed the effect of combining feature selection and resampling methods to overcome the class imbalance problem for predicting preterm labor by EHG. We assessed undersampling, oversampling, and hybrid methods applied to the training and validation dataset during feature selection by genetic algorithm, and analyzed the resampling effect on training data after obtaining the optimized feature subset. The best strategy consisted of undersampling the majority class of the validation dataset to 1:1 during feature selection, without subsequent resampling of the training data, achieving an AUC of 94.5 ± 4.6%, average precision of 84.5 ± 11.7%, maximum F1-score of 79.6 ± 13.8%, and recall of 89.8 ± 12.1%. Our results outperformed the techniques currently used in clinical practice, suggesting the EHG could be used to predict preterm labor in clinics.

Use of logistic models to evaluate the response of superovulation treatment and embryo production in Santa Inês ewes.

The study aimed to verify the influence of the FecGE mutation in superovulated ewes and to evaluate the probability of logistic models to determine the response capacity of these ewes to superovulatory treatment. Santa Inês ewes (n = 29) were genotyped for the FecGE mutation and separated for their genotype group in carriers of the mutant E allele (FecGE/E, FecG+/E) and non-carrier (FecG+/+) alleles. The ewes underwent hormonal treatment for superovulation. Aside from the genotypes, variables included in the statistical model were reproductive status (empty, early lactation, or late lactation), age (> or < 6 years), and number of births (nulliparous, primiparous, multiparous). The carriers of the mutation could be discriminated from the non-carriers based on the number of corpora lutea, rate of frozen embryos, and fecundity. Recovery rate was significantly higher (P < 0.05) in FecGE/E (94.31%) compared to FecG+/E (63.15%) and FecG+/+ (61.90%) (P < 0.05), whereas fecundity rate of FecG+/+ ewes (50.76%) was significantly higher than FecG+/E (18.96%) and FecGE/E (32.53%) (P < 0.05). We determined in this study that the response to superovulation and embryo production can be discriminated between FecGE/E and FecG+/E ewes in relation to the FecG+/+ genotype. Logistic models that included reproductive status and mutation, or reproductive status and age, or reproductive status and number of births were effective in predicting the response to superovulatory treatment.

Triflusal in Patients with Acute Coronary Syndrome and Acetylsalicylic Acid Hypersensitivity.

BACKGROUND: Acetylsalicylic acid hypersensitivity (ASAH) limits therapeutic options in patients with acute coronary syndrome (ACS), who benefit from dual antiplatelet therapy (DAPT), especially when undergoing stent implantation. Our aim was to evaluate the safety and efficacy of triflusal in patients with ACS and ASAH. METHODS AND RESULTS: Two-center retrospective study of patients diagnosed with ACS and ASAH from January 1, 2000, to May 1, 2020. Sixty-six patients were treated with triflusal. ASAH was confirmed with tests in 15 patients (22.7%). Forty-nine patients (74.2%) presented history of other drug allergies. Fifty-nine patients (89.4%) underwent stent implantation. DAPT was prescribed for ≥12 months in 54 patients. No adverse reactions to triflusal were reported. During a median follow-up of 5.12 years [IQR 2.7-9.9], rate of cardiovascular (CV) mortality was 6.1%, nonfatal myocardial infarction 12.1%, and ischemic stroke 4.5%. No cases of definite stent thrombosis occurred. Bleeding Academic Research Consortium grade ≥2 was observed in 3 patients during follow-up. CONCLUSION: In this series of patients presenting with ACS and ASA hypersensitivity, triflusal showed good tolerability and was associated with a low rate of CV and bleeding events.

Evaluation of quality and gene expression of goat embryos produced in vivo and in vitro after cryopreservation.

In the present study, we aimed to identify morphological and molecular changes of in vivo and in vitro-produced goat embryos submitted to cryopreservation. In vivo embryos were recovered by transcervical technique from superovulated goats, whereas in vitro produced embryos were produced from ovaries collected at a slaughterhouse. Embryos were frozen by two-steps slow freezing method, which is defined as freezing to -32 °C followed by transfer to liquid nitrogen. Morphological evaluation of embryos was carried out by assessing blastocoel re-expansion rate and the total number of blastomeres. The expression profile of candidate genes related to thermal and oxidative stress, apoptosis, epigenetic, and implantation control was measured using RT-qPCR based SYBR Green system. In silico analyses were performed to identify conserved genes in goat species and protein-protein interaction networks were created. In vivo-produced embryos showed greater blastocoel re-expansion and more blastomere cells (P < 0.05). The expression level of CTP2 and HSP90 genes from in vitro cryopreserved embryos was higher than their in vivo counterparts. Unlikely, no significant difference was observed in the transcription level of SOD gene between groups. The high similarity of CPT2 and HSP90 proteins to their orthologs among mammals indicates that they share conserved functions. In summary, cryopreservation negatively affects the morphology and viability of goat embryos produced in vitro and changes the CPT2 and HSP90 gene expression likely in response to the in vitro production process.

Porcine oocyte preincubation in oviductal fluid flush before in vitro fertilization in the presence of oviductal epithelial cells improves monospermic zygote production.

The present study was designed to evaluate the effect of the combination of oviduct fluid flush (OFF) and oviduct epithelial cells (OEC) in modulating the incidence of polyspermy in pigs. Therefore, for in vitro fertilization (IVF), oocyte and sperm were co-cultured in Tris-buffered medium (TBM) either supplemented with 10% OFF (OFFD group), or in the presence of a bovine OEC monolayer (OEC group), or the oocytes were exposed to OFF for 30 min before IVF (OFFB group), or in the presence of an OEC monolayer (OFFB + OEC group). Regardless of sperm concentration used (0.5, 1.5, and 4.5 × 105 cells/ml), supplementation of IVF medium with 10% OFF led to an increased (P < 0.05) monospermy rate, without alteration (P > 0.05) of the penetration rate in comparison with the control and OEC groups. When the IVF medium was supplemented with heparin, an overall increase (P < 0.05) of the final output of the IVF system in terms of zygotes with two pronuclei (2PN) was observed in the OFFD group, compared with the control and OEC groups, at a sperm concentration of 4.5 × 105 cells/ml. At this concentration, OFFB improved the monospermy rate but decreased the penetration rate, resulting in low efficiency of monospermic zygotes production. Despite this, no major effect was observed in the developmental competence of the presumed zygotes up to the blastocyst stage. The combination of OFFB with OEC improved the penetration rate, while maintaining the high monospermic rate induced by OFFB. In conclusion, the combination of treatment of oocytes by diluted OFF 30 min before IVF, followed by IVF in the presence of OEC, improved monospermic zygote production without reducing the penetration rate, when the IVF medium was supplemented with heparin.