Protective effect of Inonotus obliquus polysaccharide on mice infected with Neospora caninum.

The study aimed to explore the protective effects of Inonotus obliquus polysaccharide (IOP) on Neospora caninum (N. caninum) infection. Our data showed that the survival rate of the mice was the highest and the survival time was the longest when the IOP was 2 mg/10 g. In agreement with these observations, IOP alleviated the pathological damage in the various organs and tissues of the mice. Compared with that in the Neosporidium infection model group, the content of N. caninum in the heart, liver, spleen, lung, kidney and brain, determined through HE staining, was significantly lower. In addition, IOP inhibited the levels of immunoglobulin G1 (IgG1) and immunoglobulin G2 (IgG2a) from the 21st to 42nd day of the administration group, whereas the levels of interleukin-12 (IL-12) and serum tumor necrosis factor alpha (TNF-α) were down-regulated at 7 d - 42 d. The production of CD4+ T lymphocytes was promoted, the number of CD8+ T lymphocytes were significantly lower and the CD4+/CD8+ ratio was significantly elevated. Furthermore, IOP effectively balanced the levels of hormones including gonadotropin-releasing hormone (GnRH), luteotropic hormone (LH) and testosterone (T) in male mice, and progesterone (PROG), estradiol (E2) and prolactin (PRL) in female mice. These findings demonstrate that IOP exerts protective effects against pathological damage caused by N. caninum infection in mice, and improve the immune function of the organism and regulate the secretion balance of sex hormones.

Synthesis of Nucleotides Bearing the 2'-O-Trifluoromethyl Group and Their Application in RNA Analogs Preparation.

The integration of fluorine atoms into biologically active organic compounds has proved to be a vital technique in small molecule drugs. This technique can substantially enhance crucial properties, including metabolic stability, lipophilicity, and bioavailability, often with a mere addition of a single fluorine atom or a trifluoromethyl group. Over the past few decades, this concept has also been applied in nucleic acid chemistry. A commonly employed 2'-OH substitution is the introduction of a 2'-deoxy-2'-fluoro (2'-F) group. The strong electronegativity of fluorine prompts the modified siRNA to readily adopt a C3'-endo conformation, resulting in significant advantages in terms of binding affinity. To enrich the toolbox of chemical modification of oligonucleotides, the replacement of the 2'-OH with the 2'-O-trifluoromethyl group has been developed in RNA analog synthesis. Oligodeoxynucleotides containing the 2'-O-trifluoromethyl group can greatly increase the thermal stability of DNA/RNA duplexes depending on the position and amount of the modification. Moreover, 2'-O-trifluoromethylated oligodeoxynucleotide also exhibited a slightly higher resistance to snake venom phosphodiesterase than the unmodified oligodeoxynucleotide. The 2'-O-trifluoromethylated oligonucleotides can emerge as a label to study RNA structure and function as well, or to develop DNA/RNA-based diagnostics. Hence, it is necessary to report an effective method for the synthesis, deprotection, purification, and characterization of oligonucleotides bearing a 2'-O-trifluoromethyl group. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Preparation of 6-N-benzoyl-5'-O-dimethoxytrityl-2'-O-trifluoromethyl adenosine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 2: Preparation of 4-N-acetyl-5'-O-dimethoxytrityl-2'-O-trifluoromethyl cytidine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 3: Preparation of 2-N-isobutyryl-5'-O-dimethoxytrityl-2'-O-trifluoromethyl guanine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 4: Preparation of 5'-O-dimethoxytrityl-2'-O-2-trifluoromethyl uridine 3'-(2-cyanoethyl N,N-diisopropyl) phosphoramidite Basic Protocol 5: Solid-phase synthesis of 2'-O-trifluoromethylated RNA analogs Basic Protocol 6: Deprotection and purification of 2'-O-trifluoromethyl-RNAs.

Detecting Eating Episodes From Wrist Motion Using Daily Pattern Analysis.

This paper presents new methods to detect eating from wrist motion. Our main novelty is that we analyze a full day of wrist motion data as a single sample so that the detection of eating occurrences can benefit from diurnal context. We develop a two-stage framework to facilitate a feasible full-day analysis. The first-stage model calculates local probabilities of eating P(Ew) within windows of data, and the second-stage model calculates enhanced probabilities of eating P(Ed) by treating all P(Ew) within a single day as one sample. The framework also incorporates an augmentation technique, which involves the iterative retraining of the first-stage model. This allows us to generate a sufficient number of day-length samples from datasets of limited size. We test our methods on the publicly available Clemson All-Day (CAD) dataset and FreeFIC dataset, and find that the inclusion of day-length analysis substantially improves accuracy in detecting eating episodes. We also benchmark our results against several state-of-the-art methods. Our approach achieved an eating episode true positive rate (TPR) of 89% with 1.4 false positives per true positive (FP/TP), and a time weighted accuracy of 84%, which are the highest accuracies reported on the CAD dataset. Our results show that the daily pattern classifier substantially improves meal detections and in particular reduces transient false detections that tend to occur when relying on shorter windows to look for individual ingestion or consumption events.

A Convenient Method for the Synthesis of 2'-O-Cyanoethylated Nucleotides and Their Application in the Solid-Phase Synthesis of Related RNA Analogs.

Although small interfering RNA (siRNA) is a key player among gene inhibition therapeutics, there are many obstacles to the development of siRNA drugs due to inherent properties of oligonucleotides, including the unsatisfactory stability of unmodified siRNA, poor pharmacokinetic distribution, and the toxicity induced by off-target effects. To maximize treatment potency, chemical modification of siRNA has undoubtedly been the most successful strategy by far. Widely applied modifications include phosphorothioate linkages, 2'-O-methyl modifications, and 2'-fluoro modifications, among others. To extend the family of chemical modifications for oligonucleotides, 2'-O-cyanoethylated RNA analogs were developed through the replacement of the 2'-hydroxyl group with a 2'-O-cyanoethyl group (-OCH2 CH2 CN). This modification can provide several advantages over unmodified RNA, such as increased stability, improved binding affinity to complementary DNA or RNA strands, and resistance to degradation by cellular nucleases. The 2'-O-cyanoethyl-modified RNAs not only are applied in RNA silencing machinery but also act as research tools for studying RNA structure and function or for developing RNA-based diagnostics. Therefore, the efficient synthesis, deprotection, purification, and characterization of 2'-O-cyanoethylated RNAs deserves more attention. This protocol describes the chemical synthesis of 2'-O-cyanoethylated nucleotides and the solid-phase synthesis, deprotection, and purification of 2'-O-cyanoethylated RNAs. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Preparation of 6-N-dimethylformamidyl-5'-O-dimethoxytrityl-2'-O-cyanoethyl adenosine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 2: Preparation of 4-N-acetyl-5'-O-dimethoxytrityl-2'-O-cyanoethyl cytidine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 3: Preparation of 2-N-dimethylformamidyl-5'-O-dimethoxytrityl-2'-O-cyanoethyl guanine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 4: Preparation of 5'-O-dimethoxytrityl-2'-O-2-cyanoethyl uridine 3'-(2-cyanoethyl N,N-diisopropyl)phosphoramidite Basic Protocol 5: Solid-phase synthesis of 2'-O-cyanoethylated RNA analogs Basic Protocol 6: Deprotection and purification of synthesized 2'-O-cyanoethyl-RNAs.

Fuzzy Attention Neural Network to Tackle Discontinuity in Airway Segmentation.

Airway segmentation is crucial for the examination, diagnosis, and prognosis of lung diseases, while its manual delineation is unduly burdensome. To alleviate this time-consuming and potentially subjective manual procedure, researchers have proposed methods to automatically segment airways from computerized tomography (CT) images. However, some small-sized airway branches (e.g., bronchus and terminal bronchioles) significantly aggravate the difficulty of automatic segmentation by machine learning models. In particular, the variance of voxel values and the severe data imbalance in airway branches make the computational module prone to discontinuous and false-negative predictions, especially for cohorts with different lung diseases. The attention mechanism has shown the capacity to segment complex structures, while fuzzy logic can reduce the uncertainty in feature representations. Therefore, the integration of deep attention networks and fuzzy theory, given by the fuzzy attention layer, should be an escalated solution for better generalization and robustness. This article presents an efficient method for airway segmentation, comprising a novel fuzzy attention neural network (FANN) and a comprehensive loss function to enhance the spatial continuity of airway segmentation. The deep fuzzy set is formulated by a set of voxels in the feature map and a learnable Gaussian membership function. Different from the existing attention mechanism, the proposed channel-specific fuzzy attention addresses the issue of heterogeneous features in different channels. Furthermore, a novel evaluation metric is proposed to assess both the continuity and completeness of airway structures. The efficiency, generalization, and robustness of the proposed method have been proved by training on normal lung disease while testing on datasets of lung cancer, COVID-19, and pulmonary fibrosis.

Clinical Practices Following Train-The-Trainer Trauma Course Completion in Uganda: A Parallel-Convergent Mixed-Methods Study.

BACKGROUND: Despite the growth of trauma training courses worldwide, evidence for their impact on clinical practice in low- and middle-income countries (LMICs) is sparse. We investigated trauma practices by trained providers in Uganda using clinical observation, surveys, and interviews. METHODS: Ugandan providers participated in the Kampala Advanced Trauma Course (KATC) from 2018 to 2019. Between July and September of 2019, we directly evaluated guideline-concordant behaviors in KATC-exposed facilities using a structured real-time observation tool. We conducted 27 semi-structured interviews with course-trained providers to elucidate experiences of trauma care and factors that impact adoption of guideline-concordant behaviors. We assessed perceptions of trauma resource availability through a validated survey. RESULTS: Of 23 resuscitations, 83% were managed without course-trained providers. Frontline providers inconsistently performed universally applicable assessments: pulse checks (61%), pulse oximetry (39%), lung auscultation (52%), blood pressure (65%), pupil examination (52%). We did not observe skill transference between trained and untrained providers. In interviews, respondents found KATC personally transformative but not sufficient for facility-wide improvement due to issues with retention, lack of trained peers, and resource shortages. Resource perception surveys similarly demonstrated profound resource shortages and variation across facilities. CONCLUSIONS: Trained providers view short-term trauma training interventions positively, but these courses may lack long-term impact due to barriers to adopting best practices. Trauma courses should include more frontline providers, target skill transference and retention, and increase the proportion of trained providers at each facility to promote communities of practice. Essential supplies and infrastructure in facilities must be consistent for providers to practice what they have learned.

A preliminary study on placental damage associated to experimental neosporosis in BALB/c mice.

Neospora caninum is a protozoan parasite which can infect a range of animals, including dogs, cattle, and sheep. Bovine neosporosis, which mainly causes abortion in cattle, results in substantial economic losses worldwide. To study the effects of N. caninum infection on the placenta, a pregnant mouse model for N. caninum infection was established. The litter size (8.6 ± 1.5) and the number of live pups (6.4 ± 1.8) of infected dams were significantly lower compared with those of non-infected dams. Trophoblast cell shrinkage and a large number of apoptosomes were detected in the placentas of the infected group. The parasite load in the placental tissue was significantly higher with time after infection. Likewise, apoptosis of placental trophoblast cells significantly increased with time after infection. Among the 66 apoptotic genes detected in this study, eight genes, including Bcl-2, were significantly differentially expressed by about > tenfold in infected and uninfected mice. The expression of BAX and tumor necrosis factor-alpha (TNF-α) was upregulated in the placental cells of the infected mice, whereas the expression of BCL-2 was downregulated. Enzyme-linked immunosorbent assays (ELISAs) showed that apoptotic protease caspase-3 level was significantly increased in placental cell suspension, and insulin-like growth factor (IGF)-2 level was significantly reduced. Acetylcholine (ACH) and placental prolactin (PL) levels were initially decreased but eventually increased. In summary, infection of mice with N. caninum caused apoptotic damage to the placental tissues, cells, and genes and affected the normal physiological functions of placenta, which may largely explain the adverse pregnancy outcomes caused by N. caninum infection in mice.

Myocardial Iron Deficiency and Mitochondrial Dysfunction in Advanced Heart Failure in Humans.

Background Myocardial iron deficiency (MID) in heart failure (HF) remains largely unexplored. We aim to establish defining criterion for MID, evaluate its pathophysiological role, and evaluate the applicability of monitoring it non-invasively in human explanted hearts. Methods and Results Biventricular tissue iron levels were measured in both failing (n=138) and non-failing control (NFC, n=46) explanted human hearts. Clinical phenotyping was complemented with comprehensive assessment of myocardial remodeling and mitochondrial functional profiles, including metabolic and oxidative stress. Myocardial iron status was further investigated by cardiac magnetic resonance imaging. Myocardial iron content in the left ventricle was lower in HF versus NFC (121.4 [88.1-150.3] versus 137.4 [109.2-165.9] µg/g dry weight), which was absent in the right ventricle. With a priori cutoff of 86.1 µg/g d.w. in left ventricle, we identified 23% of HF patients with MID (HF-MID) associated with higher NYHA class and worsened left ventricle function. Respiratory chain and Krebs cycle enzymatic activities were suppressed and strongly correlated with depleted iron stores in HF-MID hearts. Defenses against oxidative stress were severely impaired in association with worsened adverse remodeling in iron-deficient hearts. Mechanistically, iron uptake pathways were impeded in HF-MID including decreased translocation to the sarcolemma, while transmembrane fraction of ferroportin positively correlated with MID. Cardiac magnetic resonance with T2* effectively captured myocardial iron levels in failing hearts. Conclusions MID is highly prevalent in advanced human HF and exacerbates pathological remodeling in HF driven primarily by dysfunctional mitochondria and increased oxidative stress in the left ventricle. Cardiac magnetic resonance demonstrates clinical potential to non-invasively monitor MID.

Splenic Abscess in the New Millennium: A Descriptive, Retrospective Case Series.

Background: Splenic abscess is a rare infection often resulting from hematogenous spread. Immunocompromised states are commonly comorbid, and the microbiology is heterogeneous. Methods: We conducted a retrospective review of 33 cases identified by convenience sampling. Cases were treated in our institution's hospital system between May 2012 and February 2021 and classified as proven or probable based on predetermined criteria. Results: The median age was 57 years, and 58% were men. Common underlying diseases included diabetes mellitus (30%), pancreatic disease (30%), and hematological malignancy (15%). The most common mechanism of pathogenesis was hematogenous spread (n = 13). Escherichia coli, enterococcal spp., and anaerobes were frequently implicated. One case was discovered at autopsy and excluded from subsequent analyses. The median duration of antimicrobial therapy (range) was 45 (5-525) days, and the median length of index hospitalization was 20 days. Percutaneous drainage by interventional radiology was common (17 of 32; 53%), and 6 patients underwent splenectomy. Treatment success was achieved in 14 of 32 cases (44%), with clinical stability in 3 of 32 cases (9%). Failures occurred in 13 of 32 (41%) cases, 2 of whom died from splenic abscesses. Two patients (2 of 32) were lost to follow-up. Conclusions: To our knowledge, this is the largest North American series since the turn of the century and the first to distinguish between proven and probable cases. As reflected in our series, patients with splenic abscess may require prolonged hospitalizations and courses of antimicrobial therapy. Improvements in management are needed.