Design of a multi-epitope-based vaccine candidate against Bovine Genital Campylobacteriosis using a reverse vaccinology approach.

BACKGROUND: Bovine Genital Campylobacteriosis (BGC), a worldwide distributed venereal disease caused by Campylobacter fetus subsp. venerealis (Cfv), has a relevant negative economic impact in cattle herds. The control of BGC is hampered by the inexistence of globally available effective vaccines. The present in silico study aimed to develop a multi-epitope vaccine candidate against Cfv through reverse vaccinology. RESULTS: The analysis of Cfv strain NCTC 10354 proteome allowed the identification of 9 proteins suitable for vaccine development. From these, an outer membrane protein, OmpA, and a flagellar protein, FliK, were selected for prediction of B-cell and T-cell epitopes. The top-ranked epitopes conservancy was assessed in 31 Cfv strains. The selected epitopes were integrated to form a multi-epitope fragment of 241 amino acids, which included 2 epitopes from OmpA and 13 epitopes from FliK linked by GPGPG linkers and connected to the cholera toxin subunit B by an EAAAK linker. The vaccine candidate was predicted to be antigenic, non-toxic, non-allergenic, and soluble upon overexpression. The protein structure was predicted and optimized, and the sequence was successfully cloned in silico into a plasmid vector. Additionally, immunological simulations demonstrated the vaccine candidate's ability to stimulate an immune response. CONCLUSIONS: This study developed a novel vaccine candidate suitable for further in vitro and in vivo experimental validation, which may become a useful tool for the control of BGC.

A Case of Success: Complete Response to Radium-223 in Metastatic Castration-Resistant Prostate Cancer.

Radium-223 dichloride (Ra223) is the first targeted alpha agent approved for treating metastatic castration-resistant prostate cancer (mCRPC) with bone-exclusive disease. A benefit in overall survival and time to the first symptomatic skeletal-related event was shown in the Alpharadin in Symptomatic Prostate Cancer Patients (ALSYMPCA) trial. However, this trial did not describe a bone scan response to Ra223, and there is no universal consensus about how it should be monitored. Furthermore, a scintigraphy flare phenomenon may lead to false-positive tracer uptake in responsive cases, thereby misleading the interpretation of imaging results.  We present the case of a 67-year-old male with mCRPC and exclusive bone disease treated with Ra223. The bone scintigraphy after the end of the treatment showed an apparent aggravation of the lesions, corresponding to a flare phenomenon, with an almost complete resolution after three months. The patient maintained a scintigraphic response for seven months.

Evening but not morning aerobic training improves sympathetic activity and baroreflex sensitivity in elderly patients with treated hypertension.

The blood pressure-lowering effect of aerobic training is preceded by improving cardiovascular autonomic control. We previously demonstrated that aerobic training conducted in the evening (ET) induces a greater decrease in blood pressure than morning training (MT). To study whether the greater blood pressure decrease after ET occurs through better cardiovascular autonomic regulation, this study aimed to compare MT versus ET on muscle sympathetic nerve activity (MSNA) and baroreflex sensitivity (BRS) in treated patients with hypertension. Elderly patients treated for hypertension were randomly allocated into MT (n = 12, 07.00-10.00 h) or ET (n = 11, 17.00-20.00 h) groups. Both groups trained for 10 weeks, 3 times/week, cycling for 45 min at moderate intensity. Beat-to-beat blood pressure (finger photoplethysmography), heart rate (electrocardiography) and MSNA (microneurography) were assessed at the initial and final phases of the study at baseline and during sequential bolus infusions of sodium nitroprusside and phenylephrine (modified-Oxford technique) to evaluate cardiac and sympathetic BRS. Mean blood pressure decreased significantly after ET but not after MT (-9 ± 11 vs. -1 ± 8 mmHg, P = 0.042). MSNA decreased significantly only after ET with no change after MT (-12 ± 5 vs. -3 ± 7 bursts/100 heart beats, P = 0.013). Sympathetic BRS improved after ET but not after MT (-0.8 ± 0.7 vs. 0.0 ± 0.8 bursts/100 heart beats/mmHg, P = 0.052). Cardiac BRS improved similarly in both groups (ET: +1.7 ± 1.8 vs. MT: +1.4 ± 1.9 ms/mmHg, Pphase  ≤ 0.001). In elderly patients treated for hypertension, only ET decreased mean blood pressure and MSNA and improved sympathetic BRS. These findings revealed that the sympathetic nervous system has a key role in ET's superiority to MT in blood pressure-lowering effect. KEY POINTS: Reducing muscle nerve sympathetic activity and increasing sympathetic baroreflex sensitivity plays a key role in promoting the greater blood pressure reduction observed with evening training. These findings indicated that simply changing the timing of exercise training may offer additional benefits beyond antihypertensive medications, such as protection against sympathetic overdrive and loss of baroreflex sensitivity, independent markers of mortality. Our new findings also suggest new avenues of investigation, such as the possibility that evening aerobic training may be beneficial in other clinical conditions with sympathetic overdrive, such as congestive heart failure and hypertrophic cardiomyopathy.

Carbamoylase-based impedimetric electronic tongue for rapid detection of paralytic shellfish toxins.

Phytotoxins produced by marine microalgae, such as paralytic shellfish toxins (PSTs), can accumulate in bivalve molluscs, representing a human health concern due to the life-threatening symptoms they cause. To avoid the commercialization of contaminated bivalves, monitoring programs were established in the EU. The purpose of this work is the implementation of a PST transforming enzyme-carbamoylase-in an impedimetric test for rapid simultaneous detection of several carbamate and N-sulfocarbamoyl PSTs. Carbamoylase hydrolyses carbamate and sulfocarbamoyl toxins, which may account for up to 90% of bivalve toxicity related to PSTs. Conformational changes of carbamoylase accompanying enzymatic reactions were probed by Fourier transform mid-infrared spectroscopy (FT-MIR) and electrochemical impedance spectroscopy (EIS). Furthermore, a combination of EIS with a metal electrode and a carbamoylase-based assay was employed to harness changes in the enzyme conformation and adsorption on the electrode surface during the enzymatic reaction as an analytical signal. After optimization of the working conditions, the developed impedimetric e-tongue could quantify N-sulfocarbamoyl toxins with a detection limit of 0.1 µM. The developed e-tongue allows the detection of these toxins at concentration levels observed in bivalves with PST toxicity close to the regulatory limit. The quantification of a sum of N-sulfocarbamoyl PSTs in naturally contaminated mussel extracts using the developed impedimetric e-tongue has been demonstrated.

Diagnostic delays in breast cancer among young women: An emphasis on healthcare providers.

Despite advances in breast cancer care, breast cancer in young women (BCYW) faces unique challenges, diagnostic delays, and limited awareness in many countries. Here, we discuss the challenges and consequences associated with the delayed diagnosis of BCYW. The consequences of delayed diagnosis in young women - which generally varies among developed, developing, or underdeveloped countries - are severe due to a faster breast tumor growth rate than tumors in older women, also contributing to advanced cancer stages and poorer outcomes. Though there are many underlying reasons for diagnostic delays due to age, the article delves explicitly deep into the diagnostic delay of BCYW, focusing on healthcare providers, potential contributing factors, its consequences, and the urgent need to start minimizing such incidences. The article suggests several strategies to address these issues, including increasing awareness, developing educational programs for healthcare providers to identify signs and symptoms in young women, developing clear diagnostic guidelines, and improving screening strategies.

Leg dystrophic calcification as a consequence of chronic diabetic foot infection: a case report.

Foot ulceration and infection is associated with a substantial increase in morbidity and mortality in patients with diabetes. We present a clinical case of recurrent diabetic foot infection with an atypical clinical evolution. A 58-year-old male patient with type 1 diabetes and a history of bilateral Charcot foot neuroarthropathy was followed at our Diabetic Foot Clinic for an unhealed plantar foot ulcer for >1.5 years with recurrent episodes of infection. He was admitted to hospital due to foot ulcer reinfection with sepsis and ipsilateral lower limb cellulitis. The foot infection was found to be associated with an underlying abscess in the anterior compartment of the leg, with a cutaneous fistulous course with extensive alterations of an inflammatory nature. Exudate from the lesion was drained and tissue biopsied, revealing Serratia marcescens and Klebsiella oxytoca with dystrophic calcification (DC). Surgical excision of dystrophic tissue with debridement of the fistulous tracts was performed. The excised material corroborated the presence of fibroadipose connective tissue with marked DC, as well as areas of mixed inflammation compatible with a chronic infectious aetiology. Targeted long-term antibiotic therapy was implemented, for a total of six weeks, with a favourable clinical evolution and complete closure of the lesion at the final follow-up. DC results from calcium deposition in degenerated tissues without evidence of systemic mineral imbalance and is a potential cause of non-healing ulcers. Few cases of DC have been reported in diabetic foot patients and its treatment remains challenging and controversial. A longer follow-up period is necessary to verify the effectiveness of our approach.

A cross-sectional study to evaluate utility measure and health-related quality of life (HRQoL) among patients with severe uncontrolled asthma in Spain.

BACKGROUND AND PURPOSE OF THE STUDY: The utility measure is a method to quantify health-related quality of life according to the preference values that patients attach to their health status. This study aimed to estimate the utility measure of patients with controlled and uncontrolled severe asthma (SA) in Spain, separately. Additionally, other characteristics (sociodemographic, clinical, and healthcare resource use [HCRU]) were also assessed for both SA populations. METHODS: This cross-sectional study included 159 patients with SA in Spain. Data were collected from medical records and directly from the patients during the study visit. Asthma Control Questionnaire (ACQ)-5 was used to classify patients with controlled and uncontrolled SA. RESULTS: Most of the patients were female (72.0% uncontrolled SA and 63.6% controlled SA). The mean (SD) EuroQol-5D (EQ-5D-5L) score was 0.88 (0.14) and 0.70 (0.25) in controlled and uncontrolled SA, respectively. The mean (SD) Asthma Quality-of-Life-5D (AQL-5D) score was 0.93 (0.09) and 0.85 (0.09) in controlled and uncontrolled SA, respectively. Emergency visits (19.2 vs. 2.7%) and hospitalizations (7.7% vs. no hospitalization) were more common among uncontrolled SA than controlled SA. Mean (SD) number of visits to primary care and pneumologists in uncontrolled SA vs. controlled SA was 4.1 (2.8) vs. 2.5 (3.0) and 3.7 (3.5) vs. 2.8 (2.2), respectively. CONCLUSION: The study provides data on utility measures among patients with SA in Spain for the first time. Patients with uncontrolled SA had lower HRQoL and higher HCRU than patients with controlled SA. Therefore, the implementation of measures that improve HRQoL among patients with uncontrolled SA is highly recommended.

Despite the existence in Spain of validated asthma questionnaires, the impact of severe asthma on quality of life, depending on whether it is controlled or not, had never been assessed.This study, which included 159 patients, was conducted to fill the gap above by obtaining two utility measures for quality of life, a generic one using the EQ-5D questionnaire (which can be used for comparison with other chronic conditions) and an asthma-specific one using the AQL-5D questionnaire.Patients with uncontrolled SA had a lower utility measure than patients with controlled disease and, therefore, a lower quality of life. In addition, patients with uncontrolled SA also had higher use of healthcare resources.These results highlight that the implementation of measures that improve the quality of life among patients with uncontrolled SA is highly recommended.

Evaluating the effects of moonlight on the vertical flight profiles of three western palaearctic swifts.

Recent studies have suggested the presence of moonlight mediated behaviour in avian aerial insectivores, such as swifts. Here, we use the combined analysis of state-of-the-art activity logger data across three swift species, the common, pallid and alpine swifts, to quantify flight height and activity in responses to moonlight-driven crepuscular and nocturnal light conditions. Our results show a significant response in flight heights to moonlight illuminance for common and pallid swifts, i.e. when moon illuminance increased flight height also increased, while a moonlight-driven response is absent in alpine swifts. We show a weak relationship between night-time illuminance-driven responses and twilight ascending behaviour, suggesting a decoupling of both crepuscular and night-time behaviour. We suggest that swifts optimize their flight behaviour to adapt to favourable night-time light conditions, driven by light-responsive and size-dependent vertical insect stratification and weather conditions.

Risk of Second Tumors in Retinoblastoma Survivors after Ionizing Radiation: A Review.

Retinoblastoma (RB) is the most common ocular neoplasm in children, whose development depends on two mutational events that occur in both alleles of the retinoblastoma susceptibility gene (RB1). Regarding the nature of these mutational events, RB can be classified as hereditary if the first event is a germline mutation and the second one is a somatic mutation in retina cells or nonhereditary if both mutational events occur in somatic cells. Although the rate of survival of RB is significantly elevated, the incidence of second malignant neoplasms (SMNs) is a concern, since SMNs are the main cause of death in these patients. Effectively, RB patients present a higher risk of SMN incidence compared to other oncology patients. Furthermore, evidence confirms that hereditary RB survivors are at a higher risk for SMNs than nonhereditary RB survivors. Over the decades, some studies have been performed to better understand this subject, evaluating the risk of the development of SMNs in RB patients. Furthermore, this risk seems to increase with the use of ionizing radiation in some therapeutic approaches commonly used in the treatment of RB. This review aims to clarify the effect of ionizing radiation in RB patients and to understand the association between the risk of SMN incidence in patients that underwent radiation therapy, especially in hereditary RB individuals.

Ultrafast mode-locked laser in nanophotonic lithium niobate.

Mode-locked lasers (MLLs) generate ultrashort pulses with peak powers substantially exceeding their average powers. However, integrated MLLs that drive ultrafast nanophotonic circuits have remained elusive because of their typically low peak powers, lack of controllability, and challenges when integrating with nanophotonic platforms. In this work, we demonstrate an electrically pumped actively MLL in nanophotonic lithium niobate based on its hybrid integration with a III-V semiconductor optical amplifier. Our MLL generates [Formula: see text]4.8-ps optical pulses around 1065 nm at a repetition rate of ∼10 GHz, with energies exceeding 2.6 pJ and peak powers beyond 0.5 W. The repetition rate and the carrier-envelope offset frequency of the output can be controlled in a wide range by using the driving frequency and the pump current, providing a route for fully stabilized on-chip frequency combs.

Empowerment-based nutrition interventions on blood pressure: a randomized comparative effectiveness trial.

Introduction: Empowerment lifestyle programs are needed to reduce the risk of hypertension. Our study compared the effectiveness of two empowerment-based approaches toward blood pressure (BP) reduction: salt reduction-specific program vs. healthy lifestyle general program. Methods: Three hundred and eleven adults (median age of 44 years, IQR 34-54 years) were randomly assigned to a salt reduction (n = 147) or a healthy lifestyle program (n = 164). The outcome measures were urinary sodium (Na+) and potassium (K+) excretion, systolic (SBP) and diastolic (DBP) blood pressure, weight, and waist circumference. Results: There were no significant differences in primary and secondary outcomes between the two program groups. When comparing each program to baseline, the program focused on salt reduction was effective in lowering BP following a 12-week intervention with a mean change of -2.5 mm Hg in SBP (95% CI, -4.1 to -0.8) and - 2.7 mm Hg in DBP (95% CI, -3.8 to -1.5) in the intention-to-treat (ITT) analysis. In the complete-case (CC) analysis, the mean change was -2.1 mm Hg in SBP (95% CI, -3.7 to -0.5) and - 2.3 mm Hg in DBP (95% CI, -3.4 to -1.1). This effect increases in subjects with high-normal BP or hypertension [SBP - 7.9 mm Hg (95% CI, -12.5 to -3.3); DBP - 7.3 mm Hg (95% CI, -10.2 to -4.4)]. The healthy lifestyle group also exhibited BP improvements after 12 weeks; however, the changes were less pronounced compared to the salt reduction group and were observed only for DBP [mean change of -1.5 mm Hg (95% CI, -2.6 to -0.4) in ITT analysis and - 1.4 mm Hg (95% CI, -2.4 to -0.3) in CC analysis, relative to baseline]. Overall, improvements in Na+/K+ ratio, weight, and Mediterranean diet adherence resulted in clinically significant SBP decreases. Importantly, BP reduction is attributed to improved dietary quality, rather than being solely linked to changes in the Na+/K+ ratio. Conclusion: Salt-focused programs are effective public health tools mainly in managing individuals at high risk of hypertension. Nevertheless, in general, empowerment-based approaches are important strategies for lowering BP, by promoting health literacy that culminates in adherence to the Mediterranean diet and weight reduction.

Visible-to-mid-IR tunable frequency comb in nanophotonics.

Optical frequency comb is an enabling technology for a multitude of applications from metrology to ranging and communications. The tremendous progress in sources of optical frequency combs has mostly been centered around the near-infrared spectral region, while many applications demand sources in the visible and mid-infrared, which have so far been challenging to achieve, especially in nanophotonics. Here, we report widely tunable frequency comb generation using optical parametric oscillators in lithium niobate nanophotonics. We demonstrate sub-picosecond frequency combs tunable beyond an octave extending from 1.5 up to 3.3 µm with femtojoule-level thresholds on a single chip. We utilize the up-conversion of the infrared combs to generate visible frequency combs reaching 620 nm on the same chip. The ultra-broadband tunability and visible-to-mid-infrared spectral coverage of our source highlight a practical and universal path for the realization of efficient frequency comb sources in nanophotonics, overcoming their spectral sparsity.

In situ extraction/encapsulation of olive leaves antioxidants in zein for improved oxidative stability of edible oils.

This study presents a sustainable and cost-effective method for preserving the bioactivity of phenolic compounds in olive leaves (OLE) during their application. The extraction and nanoencapsulation of OLE were performed in a single-step process using a rotor-stator system with zein as the encapsulating agent. The nanoprecipitation step was carried out using an aqueous sodium caseinate solution, resulting in spherical particles with an average diameter of about 640 nm, as confirmed by Transmission Electron Microscopy. Thermal characterization showed that the produced nanoparticles were more thermally stable than free OLE until 250 °C, and FTIR spectra indicated effective interaction between the phenolic compounds and zein. Antioxidant activity was evaluated using TBARS, DPPH, ABTS, and FRAP assays, with results showing that encapsulated OLE had lower antioxidant activity than free OLE. The best antioxidant capacity results were determined by TBARS assay, with IC50 results equal to 43 and 103 µgOLE/mL for free and encapsulated OLE, respectively. No anti-inflammatory potential was detected for both samples using the RAW 264.7 model, and only free OLE showed cytotoxic activity against lung cancer and gastric carcinoma. Encapsulated and free OLE were used as antioxidants in soy, palm, and palm kernel oils and compared to BHT using Rancimat. The Schaal Oven Test was also performed, and the PARAFAC chemometric method analyzed the UV-Vis spectra, which revealed high stability of the oil when 300 mg or the nanoparticles were added per kg oil. Results suggested that zein-encapsulated olive leaf antioxidants can improve the oxidative stability of edible oils.

The Evolving Pathways of the Efficacy of and Resistance to CDK4/6 Inhibitors in Breast Cancer.

The approval of cyclin-dependent kinase 4 and 6 inhibitors (CDK4/6i) in combination with endocrine therapy (ET) has remarkably improved the survival outcomes of patients with advanced hormone receptor-positive (HR+) breast cancer (BC), becoming the new standard of care treatment in these patients. Despite the efficacy of this therapeutic combination, intrinsic and acquired resistance inevitably occurs and represents a major clinical challenge. Several mechanisms associated with resistance to CDK4/6i have been identified, including both cell cycle-related and cell cycle-nonspecific mechanisms. This review discusses new insights underlying the mechanisms of action of CDK4/6i, which are more far-reaching than initially thought, and the currently available evidence of the mechanisms of resistance to CDK4/6i in BC. Finally, it highlights possible treatment strategies to improve CDK4/6i efficacy, summarizing the most relevant clinical data on novel combination therapies involving CDK4/6i.

Targeting Inhibitor of Apoptosis Proteins to Overcome Chemotherapy Resistance-A Marriage between Targeted Therapy and Cytotoxic Chemotherapy.

Precision oncology is the ultimate goal of cancer treatment, i.e., to treat cancer and only cancer, leaving all the remaining cells and tissues as intact as possible. Classical chemotherapy and radiotherapy, however, are still effective in many patients with cancer by effectively inducing apoptosis of cancer cells. Cancer cells might resist apoptosis via the anti-apoptotic effects of the inhibitor of apoptosis proteins. Recently, the inhibitors of those proteins have been developed with the goal of enhancing the cytotoxic effects of chemotherapy and radiotherapy, and one of them, xevinapant, has already demonstrated effectiveness in a phase II clinical trial. This class of drugs represents an example of synergism between classical cytotoxic chemo- and radiotherapy and new targeted therapy.

Establishment of an ultrasound-guided protocol for the assessment of hip joint osteoarthritis in rabbits-A sonoanatomic study.

Ultrasound (US) has emerged as one of the most applied imaging tools to diagnose musculoskeletal disorders and assist in guided intra-articular administrations. Nevertheless, in evaluating the rabbit hip joint, there is a need for an ultrasonographic approach. Therefore, this study aimed to describe the hip sonoanatomy, develop and validate a US-guided protocol to assess the hip joint in rabbits and apply this protocol in vivo. This study was carried out in three phases, phase I: a pilot cadaveric study, to assess the applicability of different US approaches in the hip of rabbits and, consequently, develop a detailed US-guided protocol (2 rabbit cadavers, n = 4 hips); phase II: validation of the established US-guided protocol through a numerical scoring system in healthy joints (11 rabbit cadavers, n = 22 hips), and, lastly, phase III: application of the US-guided protocol in vivo in osteoarthritic joints (5 rabbits, n = 5 hips). A total of six planes were validated, two in the ventral approach and four in the dorsal approach. The ventral transverse plane was deemed more informative regarding the hip joint sonoanatomy, enabling the identification of a greater number of structures when compared to the other planes. Nevertheless, this study suggested that the isolated application of a plane was deemed insufficient for a complete and detailed evaluation of the hip joint anatomy, rendering it necessary to employ other planes complementarily. Furthermore, the established US-guided protocol allowed a definitive diagnosis of OA, and osteophytes and capsular hypertrophy were among the defects most frequently detected. This novel study provided US anatomical landmarks for forthcoming therapeutic research and monitoring of OA development, granting the accurate identification of osseous and cartilaginous defects.

A 3D-Printed Ceramics Innovative Firing Technique: A Numerical and Experimental Study.

Additive manufacturing (AM), also known as three-dimensional (3D) printing, allows the fabrication of complex parts, which are impossible or very expensive to produce using traditional processes. That is the case for dinnerware and artworks (stoneware, porcelain and clay-based products). After the piece is formed, the greenware is fired at high temperatures so that these pieces gain its mechanical strength and aesthetics. The conventional (gas or resistive heating elements) firing usually requires long heating cycles, presently requiring around 10 h to reach temperatures as high as 1200 °C. Searching for faster processes, 3D-printed stoneware were fired using microwave (MW) radiation. The pieces were fired within 10% of the conventional processing time. The temperature were controlled using a pyrometer and monitored using Process Temperature Control Rings (PTCRs). An error of 1.25% was calculated between the PTCR (1207 ± 15 °C) and the pyrometer (1200 °C). Microwave-fast-fired pieces show similar mechanical strength to the references and to the electrically fast-fired pieces (41, 46 and 34 (N/mm2), respectively), presenting aesthetic features closer to the reference. Total porosities of ~4%, ~5% and ~9% were determined for microwave, electrically fast-fired and reference samples. Numerical studies have shown to be essential to better understand and improve the firing process using microwave radiation. In summary, microwave heating can be employed as an alternative to stoneware conventional firing methods, not compromising the quality and features of the processed pieces, and with gains in the heating time.

Immunotherapy around the Clock: Impact of Infusion Timing on Stage IV Melanoma Outcomes.

Although the impact of circadian timing on immunotherapy has yet to be integrated into clinical practice, chronoimmunotherapy is an emerging and promising field as circadian oscillations are observed in immune cell numbers as well as the expression of immunotherapy targets, e.g., programmed cell death protein-1 and its ligand programmed death ligand 1. Concurrent retrospective studies suggest that morning infusions may lead to higher effectiveness of immune checkpoint inhibitors in melanoma, non-small cell lung cancer, and kidney cancer. This paper discusses the results of a retrospective study (2016-2022) exploring the impact of infusion timing on the outcomes of all 73 patients with stage IV melanoma receiving immunotherapy at a particular medical center. While the median overall survival (OS) was 24.2 months (95% confidence interval [CI] 9.04-39.8), for a median follow-up of 15.3 months, our results show that having more than 75% of infusions in the afternoon results in shorter median OS (14.9 vs. 38.1 months; hazard ratio 0.45 [CI 0.23-0.86]; p < 0.01) with more expressive impacts on particular subgroups: women, older patients, and patients with a lower tumor burden at the outset of immunotherapy. Our findings highlight the potential benefits of follow-up validation in prospective and translational randomized studies.

Characterization of circulating microRNA profiles of postpartum dairy cows with persistent subclinical endometritis.

Subclinical endometritis (SCE) is an unresolved inflammation of the endometrium of postpartum dairy cows, seriously affecting fertility. Current diagnosis, which relies on uterine cytology or even more invasive biopsy sampling, would benefit from the identification of blood-based diagnostic biomarkers. Due to the known role of microRNAs (miRNAs) in other diseases, this case-control study evaluated the cell-free circulating miRNA profiles of SCE cows, and the network of transcripts predicted to interact with those miRNAs, previously identified as differentially expressed genes (DEG) in the endometrium of the same cows. Healthy (H, n = 6) and persistent SCE (n = 11) cows characterized by endometrial cytology and biopsy were blood sampled at 21 and 44 d postpartum (DPP). Following extraction of cell-free plasma miRNAs and RNA-seq analysis, differential abundance analysis of miRNAs was performed with the DESeq2 R package (adjusted p-value of 0.05), and in silico prediction of miRNA-interacting genes on a sequence complementary basis was conducted using the miRWalk database. The principal component analysis showed a clear clustering between groups of uterine health phenotypes (H vs. SCE), although the clustering between groups was less pronounced at 44 DPP than at 21 DPP. No effect of the stage (21 vs. 44 DPP) was observed. A total of 799 known circulating miRNAs were identified, from which 34 demonstrated differential abundance between H and SCE cows (12 less abundant and 22 more abundant in SCE than in H cows). These 34 miRNAs are predicted to interact with 10,104 transcripts, among which 43, 81, and 147 were previously identified as differentially expressed in, respectively, endometrial luminal epithelial, glandular epithelial, and stromal cells of the same cows. This accounts for approximately half of the DEG identified between those H and SCE cows, including genes involved in endometrial cell proliferation, angiogenesis and immune response, whose dysregulation in SCE cows may impair pregnancy establishment. From 219 miRNAs with mean normalized read counts above 100, the presence and abundance of miR-425-3p and miR-2285z had the highest discriminatory level to differentiate SCE from H cows. In conclusion, despite apparent confinement to the endometrium, SCE is associated with a distinct circulating miRNA profile, which may represent a link between the systemic changes associated with disease and the endometrial immune response. The validation of a miRNA panel consisting of circulating cell-free miR-425-3p and miR-2285z may prove a relevant advancement for the noninvasive diagnosis of persistent SCE.