Although microaerobic activated sludge process has been developed to convert nitrogen compounds in fermentation industrial wastewater to ammonium, its applicability at the industrially relevant situations is largely unknown. Here, responses of the treatment performances and microbial communities to the sequential settings of the low DO and/or low pH conditions for the effective start-up of the practically downscaled processes fed with the simulated wastewater derived from real streams were investigated by the 3D fluorescence spectroscopy, LC-TOF-MS, qPCR of nitrification-related functional genes and high-throughput sequencing of 16S/18S rRNA genes. The ammonium conversion and retention rates increased rapidly to 77.6-103.6% in all the settings, highlighting the relevance of the process in the context of nitrogen recovery with a combination of the downstream concentration and separation steps in practical application. The advancement was associated with the formation of the stable whole prokaryotic communities and the decline (∼2 order decreases in gene copies) of the abundant nitrifier ammonia-oxidizing bacteria (AOB). The hydrolytic and/or proteolytic Extensimonas soli and Thermomonas haemolytica commonly proliferated and predominated at ∼51.1% and ∼26.5% of the total, suggesting their central involvements in ammonification. The TOC removal rates were relatively high (>88.3%), although exhibiting the trade-off relationship with nitrogen recovery, for which some dissolved organic residues, e.g., amino acid-related substances, were identified. An issue of this process was the increases in the treated wastewater turbidities but could be alleviated by DO control. The eukaryotes Amoebozoa and Ochrophyta, as well as AOB, were responsible for the sludge flocculation, owing to the comparison with the conventional nitrification-denitrification process with the low turbidities. Consequently, this study illuminated the high adaptabilities of microbial communities to the changing conditions for establishment of the novel nitrogen circular technology.
This study investigated the feasibility of accelerating aerobic granular sludge (AGS) formation by short-term in-situ dosing of chitosan (CTS) in sequencing batch reactors. Compared with the control reactor, short-term CTS dosing during start-up markedly promoted biomass aggregation, shortened the complete granulation time from 27 to 9 days, and produced larger and denser granules with superior settleability. In the CTS-amended reactor, the mean granule size reached 752 µm, while sludge volume index at 30 min (SVI30) rapidly decreased to 59.4 mL/g within 24 h and stabilized at 38.2-44.8 mL/g during steady operation. Mechanistic analyses showed that CTS reduced sludge surface electronegativity, stimulated extracellular polymeric substance (EPS) secretion, and functioned as both a cationic bridging agent and a physical nucleation core. These roles were further supported by direct visualization using CTS-coated Fe3 O4 particles as a tracer. Temporary pH control applied to activate the cationic properties of CTS initially suppressed nitrification, but the inhibition was reversible. After recovery, the CTS reactor achieved superior total nitrogen (TN) removal, with an average effluent TN of 8.6 mg/L compared with 15.3 mg/L in the control. The enhanced TN removal may be associated with the combined effects of larger and more compact granules and the possible contribution of residual CTS as supplementary electron donor. Microbial analysis further revealed the enrichment of EPS-producing and structural bacteria, especially Thiothrix, in the CTS reactor. Overall, short-term in-situ CTS dosing provides a simple and biocompatible strategy for rapid AGS start-up and improved nitrogen removal.
Biological methane (CH4) removal from dilute air streams (200-5,000 ppm) is fundamentally constrained by slow mass transfer, primarily due to CH4's low solubility and diffusivity in water. Conventional biofilters, the current state of technology, suffer from long start-up periods, progressive pore clogging, and high transport energy requirements. This study investigates a "dry" biofilm concept designed to overcome these bottlenecks by minimizing the aqueous boundary layer while maintaining microbial activity via capillary-mediated nutrient delivery. Using concentrated biomass of the methanotroph Methylomicrobium buryatense 5GB1C, three generations of membrane-supported reactor configurations were evaluated. The third-generation (G3) design utilized a cellulose-bead capillary support to maintain a physical gap between the membrane and the liquid surface, enabling continuous drainage of metabolic water. Experimental results demonstrated that the "dry" G3 configuration achieved CH4 removal rate of 148.9 mg·m-2·hr-1 at 4000 ppm, representing a 397% improvement over the first-generation floating mesh configuration. At 500 ppm, G3 design achieved a CH4 removal rate of 18.6 mg·m-2·hr-1, corresponding to an over six-fold improvement over one of the highest reported values. Furthermore, the system enabled immediate start-up post-inoculation and maintained an optimal microenvironment pH (8.8-9.0) even as the bulk medium acidified. These results establish that replacing liquid-phase diffusion with drastically faster gas-phase transport provides a high-efficiency framework for mitigating low-concentration CH4 emissions. With the added benefits of minimal pressure drop and easy biomass harvesting via scraping, this dry biofilm approach offers a scalable and sustainable alternative for atmospheric methane mitigation.
Highly crystalline single-walled carbon nanotubes were employed as robust supports for carbon-encapsulated PtPd alloy electrocatalysts synthesized via a rapid, industrially scalable solution plasma method, enhancing long-term durability under frequent start-up/shut-down conditions. Electrochemical evaluation of PtPd@C/SWCNT as a cathode catalyst in a polymer electrolyte fuel cell (PEFC) membrane electrode assembly (MEA) demonstrated superior durability compared to commercial Pt/C and monometallic Pt@C/SWCNT under an accelerated durability test. PtPd@C/SWCNT maintained high performance for 5000 potential cycles and retained over 50% of its electrochemically active surface area (ECSA) after 10 000 cycles under the accelerated durability test of high-potential triangular pulses (1.0-1.5 V) simulating harsh conditions encountered during actual start-up/shut-down operations. Carbon encapsulation effectively inhibited nanoparticle agglomeration and suppressed the oxidation of the SWCNT support in close proximity to the nanoparticles during the durability test of 30 000 cycles. Raman spectroscopy confirmed the excellent corrosion resistance and maintained the crystallinity of the SWCNT support. The negligible thickness change observed in the PtPd@C/SWCNT cathode layer further highlights the benefit of the SWCNT support and carbon encapsulation in maintaining structural integrity under severe operating conditions. XPS analysis indicated a more stable, reduced state of Pt in PtPd@C/SWCNT compared to that of Pt/C. These results highlight the synergistic effects of the SWCNT support and carbon encapsulation in improving both catalyst stability and support durability for prolonged PEFC operation, particularly under demanding heavy-duty vehicle (HDV) conditions.
We report the mechanism underlying two-step yielding in repulsive colloidal microgel glasses under shear deformation. Strain sweep and start-up flow experiments demonstrate the existence of two-step yielding, which was further investigated by creep-recovery, and Lissajous-Bowditch curves to probe intra-cycle nonlinearities. By increasing the microgel volume fraction, we track the transition from entropic to jammed glass regimes and examine the distinct roles of particle softness and crosslinking heterogeneity in yielding behaviour. Soft core-shell particles exhibit two-step yielding in the jammed glass regime at a frequency of ω = 1 rad s-1. We compare the results for three types of particles: soft core-shell; stiff core-shell; and homogeneously crosslinked. We find that stiff core-shell and homogeneous particles do not exhibit two-step yielding under any experimental conditions. These findings demonstrate that softness combined with a core-shell particle structure is necessary to support two-step yielding. Intra-cycle nonlinearities reveal that strain stiffening develops between the first and second yield points, arising from resistance to macroscopic flow at and beyond the first G″ peak. This resistance to cage breaking originates from the strong interlocking of interpenetrated polymer chains that occurs during microgel deformation and compression in the jammed state. Macroscopic flow begins at the second yield point, where particles escape their cages by breaking the interlocking structure, leading to the G'-G″ crossover.
In passive-source seismic exploration, even after seismic instruments complete unified start-up acquisition and hardware synchronization, long-duration continuous records may still contain small residual timing errors, which in turn broaden cross-correlation peaks and degrade event-location results. To address this problem, this study proposes a wavefield-domain residual timing refinement method. The method uses stable noise windows and controlled artificial events in continuous records as constraints, and performs data-window preprocessing, reference cross-correlation function construction, pairwise residual lag estimation, confidence-weighted multi-station joint fusion, and smoothing-constrained fitting of a continuous correction curve to achieve a posterior refinement of residual timing errors after hardware synchronization. Fractional-delay interpolation is then used for waveform correction. Validation using a 60 min continuous record from a local six-station array shows that the proposed method can serve as an effective supplement to hardware synchronization, suppress residual timing errors, and improve the temporal consistency, waveform stackability, and interpretation reliability of passive-source seismic exploration data.
The REvascularization CHoices Among Under-Represented Groups Evaluation (RECHARGE) program is enrolling 1200 women, Black, and Hispanic patients in two parallel randomized trials of percutaneous coronary intervention (PCI) versus coronary artery bypass grafting (CABG). Funded by a phased Patient-Centered Outcomes Research Institute award, the pilot phase was designed to assess the feasibility of enrolling groups historically under-represented and challenging to enroll in prior revascularization trials, evaluate willingness of patients to accept randomization, refine patient and stakeholder engagement, and scale site infrastructure and data collection across diverse centers. We report key insights from the pilot phase. Physician and patient treatment preferences, often shaped by prior experience and evidence not directly applicable to these cohorts, were the main reasons eligible patients were not randomized. Many sites also lacked consistent multidisciplinary Heart Team processes for coronary disease, requiring new workflows to establish equipoise between PCI and CABG. Successful recruitment required intentional trust-building and tailored patient-facing materials, while engagement of non-academic centers demanded added financial, educational, and start-up support. During the 2-year pilot phase, 91 U.S. and 17 Canadian sites were selected, and 65 were activated. Median activation time was 10.8 months (IQR 9.1-13.6). The pilot enrollment goal of 60 participants was exceeded, with 141 patients randomized within 13 months at a mean rate of 0.27 patients/site/month, prompting expansion to up to 150 sites for the full program. The lessons learned from the pilot phase of the RECHARGE program can inform the design and implementation of future randomized trials seeking to enroll traditionally under-represented populations.
Microplastics may disturb microbial activity and biofilm development in biological wastewater treatment systems, yet the response of three-dimensional rotating biological contactor start-up biofilms to polypropylene microplastic stress remains unclear. This study evaluated a biofilm initiation strategy using heterotrophic nitrification-aerobic denitrification (HN-AD) bacteria (H-3D-RBCs) and compared it with activated sludge-inoculated systems (A-3D-RBCs) under polypropylene microplastic (PP-MP) exposure. H-3D-RBCs showed superior resistance to PP-MP disturbance, with total nitrogen removal decreasing by only 14 %, compared with an approximately 60 % decline in A-3D-RBCs. Respiratory activity inhibition remained below 15 % in H-3D-RBCs but exceeded 90 % in A-3D-RBCs. 16S rRNA gene sequencing showed that PP-MP reduced species richness and diversity in A-3D-RBCs and was associated with a > 90 % loss of core denitrifying genera, including Corynebacterium and Pseudoxanthomonas, whereas H-3D-RBCs maintained community stability and enriched Pseudoxanthomonas to 13.8 %. Metagenomic analysis indicated that PP-MP impaired nitrification and denitrification potential in A-3D-RBCs, as reflected by decreased genes encoding AMO and HAO, a 51.78 % decrease in nosZ abundance, and enhanced dissimilatory nitrate reduction to ammonium (DNRA), which likely intensified competition with denitrification and promoted nitrogen conversion to ammonia. In contrast, H-3D-RBCs suppressed DNRA and maintained high nosZ abundance. Untargeted metabolomics further showed that PP-MP was associated with metabolic disorders in A-3D-RBCs, especially disruptions in alanine, aspartate, and glutamate metabolism and arginine biosynthesis, whereas H-3D-RBCs preserved these key nitrogen metabolic processes. Overall, this study identifies key vulnerabilities of nitrogen-removal biofilms under PP-MP disturbance and provides multi-omics evidence to support the development of microplastic-resistant biofilm wastewater treatment systems.
A comprehensive review and analysis of drug discovery efforts at Indian companies between the mid-1990s and 2025 reveals 1095 ongoing (462) or past (633) projects and molecules under investigation at 195 major pharmaceutical, biotechnology, and start-up companies, of which 128 are currently actively pursuing research. They consist mainly of small molecules (811), followed by novel biologics (189) and gene therapies (95), and cover all stages from early discovery (618), preclinical (276), and clinical development phases (Phase 1: 98; Phase 2: 63; Phase 3: 19) up to approved drugs and treatments (21). Small molecules are dominated by new chemical entities (684), followed by prodrugs, salts, and formulations (56), repurposed drugs (53), and others (18). Biologics consist largely of monoclonal antibodies or fragments (92), antibody-drug conjugates (22), various (fusion) proteins (40), enzymes (10), and others (25). Gene therapies use gene silencing (24), gene transfer (21), genome editing (7), modulation of mRNA splicing (3), and genetically modified cells based on chimeric antigen receptor technology using T and NK cells (40). Tracking companies and projects over time illustrates the dynamics and increasing diversification of drug discovery activities in India.
The anterior component of force (ACF) is key to proximal contact loss (PCL) after implant restoration, yet its quantitative impact remains unclear. This study employed digital model analysis to indirectly assess the effects of ACF and examine its association with PCL. Following final restoration, 3D surface and buccal occlusal data were obtained from ninety first molar implant sites and mesial adjacent teeth. Occlusal force-induced changes in proximal contact gap (ΔdP) and centroid position (ΔdC) were measured using Geomagic Wrap 2021 for indirect ACF estimation. Proximal contact gaps were measured under non‑occluding conditions at baseline and 6 months to calculate the change over time (ΔD). Univariate and multivariate analyses identified PCL-related biomechanical influencing factors, and a restricted cubic spline (RCS) model was developed and validated using Cohen's Kappa. The 6-month PCL incidence was 17.78%, and ΔdP emerged as a significant biomechanical influencing factor of ΔD (P < 0.001). RCS analysis revealed that the adjacent state remains stable when ΔdP is within the range of - 13 μm to 9 μm; beyond this range, imbalance occurs. The model demonstrated good fitting capability (κ = 0.59, P = 0.003). This study achieved an indirect assessment and quantification of the effect of ACF through digital model analysis technology, and clarified its correlation with the PCL of the implant, providing new evidence and methodological references for related biomechanical research.
Carbon based anodes that rely solely on Li-ion storage are inherently limited in energy density, whereas systems dominated by Li-metal storage suffer from poor reversibility and short cycle life. Hybrid Li-ion/Li-metal batteries (LIB/LMBs), enabled by carbon-based Li-free anodes, offer a promising pathway by integrating Li-ion intercalation and Li-metal plating/stripping within a single framework. Despite rapid progress, the practical implementation of hybrid LIB/LMBs remains hindered by low Coulombic efficiency, unstable solid electrolyte interphase, dendrite growth, and large volume fluctuations. These challenges are fundamentally associated with irreversible Li loss and interfacial instability, which limit long-term cycling and energy efficiency. In this review, we systematically summarize recent advances in carbon-based Li-free anodes for hybrid LIB/LMBs, with a focus on (i) the hybrid storage mechanism, (ii) rational design of carbon materials, (iii) interface engineering strategies, (iv) mechanistic insights from in situ characterization, and (v) critical perspectives toward practical deployment. Particular emphasis is placed on the relationship between Coulombic efficiency, Li inventory retention, and system-level performance. This review provides a unified framework for understanding hybrid storage chemistry and offers strategic guidance for the development of next-generation high-energy-density batteries.
The genus Barringtonia (Lecythidaceae) is a vital component of tropical coastal forests and mangrove ecosystems. Among its members, B. racemosa and B. fusicarpa are classified as Endangered and Vulnerable, respectively, due to habitat degradation and anthropogenic pressures, underscoring the urgent need for genetic studies to guide conservation. Chloroplast (cp.) genomes serve as essential resources for phylogenetic reconstruction and conservation genetics. However, the scarcity of cp. genome data for Barringtonia has limited comprehensive evolutionary and conservation-oriented investigations. We assembled and annotated the first complete cp. genomes of B. racemosa, B. fusicarpa, and B. acutangula. All three genomes exhibit the typical quadripartite structure, ranging from 158,959 bp (B. racemosa) to 159,837 bp (B. acutangula), and contain 132 genes (87 protein-coding, 37 tRNA, 8 rRNA) with a GC content of 36.68%-36.86%. Collinearity and IR boundary analyses revealed high structural conservation without large-scale rearrangements. Interspecific sequence-level variations were detected in simple sequence repeats (SSRs) and long repeats. Nucleotide diversity (π) analysis identified highly polymorphic regions, including rpl20 (π = 0.080), rpoA (π = 0.064), rps3 (π = 0.063), and ndhF (π = 0.060), which represent promising molecular markers for population genetics within the genus. Codon-based selection analyses (Ka/Ks) showed that all protein-coding genes are under strong purifying selection (mean Ka/Ks 0.32-0.37), with no evidence of positive selection. Pairwise genetic distances (p-distances) among Barringtonia species are extremely low (mean 0.0046), while distances to the related genus Bertholletia are ~ 6-fold higher, supporting their generic distinction. Phylogenetic analysis robustly supports Barringtonia as a monophyletic clade (bootstrap = 100%), with B. racemosa and B. fusicarpa forming a sister lineage to B. acutangula. This study provides the first high-quality cp. genome resources for the two threatened Barringtonia species, revealing strong structural and sequence conservation but no direct chloroplast genomic correlates of endangerment. The identified polymorphic regions and repeat markers lay a foundation for future population genetics, phylogeographic studies, and conservation-oriented genetic management of these ecologically important coastal plants.
Spermiogenesis dysfunction is a major cause of male infertility; however, the underlying molecular mechanisms involved remain incompletely elucidated. Although transmembrane protein 67 (TMEM67), a ciliary transition zone protein implicated in ciliopathies, is highly enriched in mouse testes, its cell type-specific functional relevance in spermatogenesis is unclear. Here, we generated germ cell-specific (Stra8-Tmem67f/f) and Sertoli cell-specific (Amh-Tmem67f/f) Tmem67 knockout mice to investigate the function of TMEM67 in spermatogenesis and male fertility. Amh-Tmem67f/f mice maintained normal fertility and exhibited normal spermatogenesis, with no significant differences in testicular histology or sperm count, morphology, or motility compared with wild-type (WT) controls. However, Stra8-Tmem67f/f males were completely infertile, manifesting severe oligoasthenoteratozoospermia (OAT) characterized by a drastic reduction in sperm count, total loss of sperm motility, and global sperm malformation. Further investigations revealed that TMEM67 deletion did not impair spermatogonial proliferation or meiosis, but instead disrupted key spermiogenic events, including manchette dynamics, acrosome biogenesis, and flagellum development. Proteomic analysis indicated that TMEM67 knockout altered the expression of numerous spermiogenesis-related proteins. Furthermore, our experiments confirmed that TMEM67 deficiency led to profound perturbations in both the expression levels and subcellular localization of key spermiogenic regulators in the testis. Collectively, our findings demonstrate that TMEM67 is indispensable for spermiogenesis and male fertility, revealing its critical role in coordinating manchette function, axonemal integrity, and spermiogenesis-related protein regulation, providing novel insights into OAT pathogenesis.
Agricultural residues, such as corn stover, generate more than 2 billion tons of biomass annually, presenting challenges for efficient bioenergy conversion due to their lignocellulosic structure and microbial instability under suboptimal conditions. These limitations reduce methane yields and hinder system scalability in anaerobic digestion processes. This study examined the effects of temperature variations and digestate recirculation on methane production from corn stover. Batch biochemical methane potential assays were conducted at 25 °C, 37 °C, and 55 °C, followed by continuous stirred-tank reactor experiments with organic loading rates of 2.1-3.1 g VS/(L·d); microbial community analysis involved 16 S rRNA sequencing and correlation assessments. The thermophilic conditions (55 °C) resulted in methane yields of 218 mL/g VS, representing increases of 169% and 12% over the psychrophilic (25 °C) and mesophilic (37 °C) conditions; digestate recirculation in the continuous reactors reduced volatile fatty acid accumulation by 23-55% and increased yields to 162 mL/g VS at 2.6 g VS/(L·d), with shifts in the bacterial and archaeal communities favoring lignocellulose degradation and methanogenesis. These results demonstrate improved anaerobic digestion efficiency through integrated strategies, advanced bioenergy production and waste conversion by optimizing microbial stability and process kinetics.
Enhancing myeloid effector cell recruitment may improve immunotherapy by monoclonal antibodies, including those against acute lymphoblastic leukemia (ALL). To assess the expression of target antigens in B-cell ALL (B-ALL), we compared mRNA profiling of 559 patient leukemia samples across 18 molecular subtypes with that of representative cell lines. The latter served as target cells to compare human immunoglobulin G1 (IgG1) or IgA2 variants against CD19, CD20, or CD38 in antibody-dependent cellular phagocytosis (ADCP) by macrophages and antibody-dependent cell-mediated cytotoxicity (ADCC) by polymorphonuclear leukocytes (PMN). Interestingly, antibodies against broadly expressed CD19 were negligibly effective in mediating ADCP or ADCC. Antibodies against CD20 or CD38, the former variably expressed across subtypes, triggered ADCP by macrophages both as IgG1 and IgA2. However, PMN-mediated ADCC against CD20 or CD38 was only observed with IgA2 variants but not with respective IgG1 antibodies. Blocking the myeloid checkpoint CD47/signal regulatory protein α (SIRPα) enhanced ADCP and ADCC by IgA2 antibodies against CD20 and CD38 but not CD19. Both CD47 and its enzymatic modifier glutaminyl-peptide cyclotransferase-like protein (QPCTL), which can be inhibited by small molecules, were broadly expressed across B-ALL subtypes. QPCTL catalyzes the formation of an N-terminal pyroglutamic acid on CD47, which is directly involved in CD47/SIRPα interactions as shown by novel engineered CD47 variants. Importantly, the combination of anti-CD38 IgA2 and CD47 blockade was effective against xenografted B-ALL in human FcαRI (CD89) transgenic NXG mice. Together, these studies support combining anti-CD38 IgA2 with CD47 interference to improve myeloid effector cell recruitment for B-ALL immunotherapy.
The necessity of iodinated contrast agents in radiographic diagnosis comes with an increased risk of kidney injury. The sclerotium of Poria cocos (Schw.) Wolf, a traditional Chinese medicine, exhibits anti-inflammatory and diuretic pharmacological properties. Its extracted polysaccharides demonstrate anti-apoptotic, antioxidant, and anti-inflammatory activities. This study aimed to investigate the effects and potential mechanisms of Poria cocos polysaccharides (PCP) on contrast-induced acute kidney injury (CI-AKI). The purified PCP was characterized using FTIR and monosaccharide analysis. A CI-AKI mouse model was established to evaluate its preventive effects via oral gavage. Mitochondrial structure, function, and biogenesis were assessed by measuring mitochondrial membrane potential (MMP), ROS, ATP, and mitochondrial DNA copy number. Antioxidant capacity and apoptosis levels were evaluated by detecting 8-OHdG, caspase 3/7 activity, cleaved-PARP, MDA, and GSH. Finally, the mechanism of action of PCP was analyzed using qPCR and immunoblotting. The results showed that PCP significantly mitigated CI-AKI by inhibiting contrast-induced apoptosis and oxidative stress in kidney tissue. Furthermore, PCP maintained mitochondrial homeostasis in the kidneys by enhancing mitochondrial biogenesis and antioxidant capacity. These findings were consistent with in vitro data. Notably, PGC-1α was identified as a key factor in the nephroprotective effects of PCP, with its activation attributed to the CaMKK2-AMPK signaling pathway. These findings suggest that PCP offers significant benefits in mitigating CI-AKI by enhancing antioxidant defenses and promoting mitochondrial biogenesis through activation of the CaMKK2/AMPK/PGC-1α signaling pathway in the kidney. Therefore, PCP could be a potential preventive agent for CI-AKI.
Electrocatalytic hydrogen evolution reaction (HER) necessitates managing both macroscopic wettability for bubble release and microscopic water structure for proton transfer. However, highly hydrophilic surfaces, while beneficial for bubble transport, tend to induce strongly hydrogen-bonded water networks that hinder proton dynamics, presenting a fundamental conflict in advancing electrocatalyst design. Herein, we leverage polymeric modifiers to tune the surface electronic structure of platinum electrodes and investigate the synergistic influence of macroscopic wettability and microscopic interfacial water structure on HER performance. We demonstrate that peak HER performance is achieved on polyethylenimine (PEI)-modified Pt electrode, which uniquely affords moderate hydrophilicity (contact angle: ∼15°) alongside a disordered, weakly hydrogen-bonded interfacial water network. This dual optimization enhances bubble release and proton-transfer kinetics, reducing the overpotential by 229 mV at 10 mA cm-2 compared to pristine Pt. In situ Raman spectroscopy confirms an increase in weakly hydrogen-bonded water species under PEI modification compared to the superhydrophilic electrode. This study bridges interfacial water dynamics with electrode wettability, guiding the design of advanced electrocatalytic interfaces.
To provide a detailed description of the clinical procedure for aspirating small antral follicles (2.0-8.0 mm) as part of the Saigon biphasic in vitro maturation (IVM) protocol. This includes indications for the procedure, patient and equipment preparation, the aspiration process, laboratory techniques for identifying cumulus-oocyte complexes (COCs), and the timing for embryo transfer. This instructional video presents real-time footage of clinical procedures and embryology practices. It offers step-by-step guidance and practical recommendations for conducting a gonadotropin-free biphasic IVM cycle, also known as capacitation-IVM (CAPA-IVM). Patients undergoing IVM treatment. All individuals featured in the video have provided informed consent for its publication and online distribution, including on social media, journal websites, scientific platforms, and other relevant platforms. The video includes an introduction and patient consent process; indications for IVM; timing of oocyte aspiration, patient and equipment preparation, including types of aspiration needles; potential technical and clinical complications; timing for fresh embryo transfer; visual demonstration of the aspiration procedure, including ultrasound guidance, needle pressure, flushing, and aspiration technique; and laboratory demonstration of oocyte search and IVM preparation, including filtration of follicular aspirate and identification/classification of compact COCs. Clinical techniques for immature oocyte retrieval and compact COCs identification for the gonadotropin-free CAPA-IVM protocol. Numerous viable COCs can be retrieved successfully using the oocyte retrieval techniques outlined for gonadotropin-free CAPA-IVM. The video demonstrates an efficient and flexible approach to identifying compact COCs, adaptable to various clinical conditions. Embryo transfer subsequently may be performed as either a fresh or frozen cycle, depending on the patient's choice. (s): In vitro maturation is a safe and effective treatment option for eligible patients, particularly those with high ovarian reserve or polycystic ovary syndrome. This technique enables the retrieval of multiple immature COCs without ovarian stimulation. The procedures outlined may differ from those used in other centers, offering a standardized approach for collecting COCs from small follicles for IVF and gamete banking. The Saigon protocol, which combines gonadotropin-free CAPA-IVM with fresh embryo transfer, underscores the clinical potential of IVM as a practical alternative to conventional stimulation protocols.
Hepatocellular carcinoma (HCC) is a malignant tumour with high global incidence and mortality, featuring limited therapeutic options and poor prognosis, which necessitates novel prognostic biomarkers. The immune checkpoint molecule CD274 (Programmed Death-Ligand 1, PD-L1) correlates with poor prognosis in most solid tumours, yet its prognostic value in HCC remains controversial, and circulating levels are unclear. This study aimed to multi-dimensionally investigate CD274's expression patterns and clinical significance in HCC. Bioinformatics analyses were performed on The cancer genome atlas (TCGA) and Genotype-Tissue Expression (GTEx) databases to screen differentially expressed genes, with functional and pathway annotations via Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Set Enrichment Analysis (GSEA), and Single-Sample Gene Set Enrichment Analysis (ssGSEA). Survival analysis and univariate/multivariate Cox proportional hazards regression models were used to assess the prognostic value of CD274. PD-L1 protein expression was validated by immunohistochemistry (IHC) on HCC tissue microarrays. Serum PD-L1 concentrations were detected by ELISA in treatment-naive HCC patients and healthy controls. CD274 was lowly expressed in HCC tissues versus normal ones, and its expression correlated with TNF-α/NF-κB, complement, IFN-γ, adipogenesis and oxidative phosphorylation pathways, as well as lymphocyte-mediated immunity, immune globulin complexes and antigen binding. High CD274 expression negatively correlated with Th17 cells but positively with helper T cells, activated dendritic cells (aDCs), Th1 cells, T cells and macrophages via ssGSEA. Patients with high CD274 expression had longer overall survival (OS), disease-specific survival (DSS) and progression-free interval (PFI); multivariate Cox regression confirmed CD274 as an independent protective factor for DSS. PD-L1 protein expression showed no significant difference between HCC and normal liver tissues. Serum PD-L1 median was 349.6 pg/mL in HCC patients, significantly higher than 181.3 pg/mL in healthy controls. This study first identifies a unique "low tissue expression and elevated peripheral blood level" pattern of CD274 in HCC, with intratumoural high expression correlating with favourable prognosis. CD274 may exert distinct roles in local tumour and systemic immune regulation, and serum CD274 is a promising prognostic biomarker for HCC worthy of further research.
Screening for diabetic retinopathy using fundus photographs is the global standard of care but results in high false-positive referrals to evaluate diabetic macular edema (DME), placing a substantial burden on specialist eye clinics. Integrating an AI-based optical coherence tomography (AI-OCT) system into screening pathways may reduce potentially unnecessary referrals. To evaluate the diagnostic and referral performance of an AI-OCT system for DME detection within a diabetic retinopathy screening pathway in clinical settings. Stepwise evaluation conducted in Hong Kong Special Administrative Region: a prospective silent-mode validation (February 2020 to July 2023) recruiting 603 patients with diabetes at a tertiary hospital triage unit, followed by a multicenter noninferiority RCT (September 2023 to April 2025), with follow-up completed in May 2025, recruiting 276 patients with suspected DME referred from a territory-wide diabetic retinopathy screening program. RCT participants were randomized to intervention (referral for DME evaluation based on both fundus photograph-based screening reports and AI-OCT reports [n = 137]) or control (automatic referral based solely on fundus photograph-based screening reports [n = 139]) groups. The AI-OCT system incorporated image-quality assessment, DME detection, and uncertainty flagging. Study outcomes focused on referral rates under the 2 pathways; for ethical reasons, all participants ultimately underwent specialist evaluation. The primary outcome was false-positive DME referral rate, with a prespecified noninferiority margin of 20%. The secondary outcomes included sensitivity and specificity for DME detection and DME referral. In prospective silent-mode validation (mean age, 64.7 [SD, 9.4] years; 56.2% male), 86 of 1200 scans (7.2%) were identified as ungradable and 49 of 1114 gradable scans (4.4%) were classified as uncertain. The system achieved 98.8% (95% CI, 94.5%-100.0%) sensitivity and 90.7% (95% CI, 88.7%-92.4%) specificity for DME detection. In the RCT (mean age, 63.9 [SD, 10.9] years; 54.7% male), DME prevalence was similar in the intervention and control groups (30.9% vs 29.9%). The false-positive DME referral rate was 24.1% (95% CI, 14.6%-37.0%) and 69.1% (95% CI, 61.0%-76.1%), respectively (absolute difference, -45% [95% CI, -58.2% to -31.9%; P < .001 for noninferiority]; upper bound of the CI below the prespecified noninferiority margin of 20%). Sensitivity for DME referral was 100.0% (95% CI, 100.0%-100.0%) in both groups. Specificity for DME referral was 86.5% (95% CI, 79.3%-92.9%) in the intervention group and 0.0% (95% CI, 0.0%-0.0%) in the control group. No cases of DME occurred among nonreferred participants in the intervention group. Compared with standard practice, incorporation of the AI-OCT system as a secondary screening tool was noninferior with respect to false-positive referral rates and was associated with a substantial reduction in potentially unnecessary DME referrals without compromising sensitivity. Chinese Clinical Trial Registry: ChiCTR2300075087.