A Janus Microsphere Delivery System Orchestrates Immunomodulation and Osteoinduction by Fine-tuning Release Profiles.

Bone regeneration is a well-orchestrated process synergistically involving inflammation, angiogenesis, and osteogenesis. Therefore, an effective bone graft should be designed to target multiple molecular events and biological demands during the bone healing process. In this study, a biodegradable gelatin methacryloyl (GelMA)-based Janus microsphere delivery system containing calcium phosphate oligomer (CPO) and bone morphogenetic protein-2 (BMP-2) is developed based on natural biological events. The exceptional adjustability of GelMA facilitates the controlled release and on-demand application of biomolecules, and optimized delivery profiles of CPO and BMP-2 are explored. The sustained release of CPO during the initial healing stages contributes to early immunomodulation and promotes mineralization in the late stage. Meanwhile, the administration of BMP-2 at a relatively high concentration within the therapeutic range enhances the osteoinductive property. This delivery system, with fine-tuned release patterns, induces M2 macrophage polarization and creates a conducive immuno-microenvironment, which in turn facilitates effective bone regeneration in vivo. Collectively, this study proposes a bottom-up concept, aiming to develop a user-friendly and easily controlled delivery system targeting individual biological events, which may offer a new perspective on developing function-optimized biomaterials for clinical use.

A Multiplex Recombinase-Aided qPCR Assay for Highly Sensitive and Rapid Detection of khe, blaKPC -2, and blaNDM -1 Genes in Klebsiella pneumoniae.

OBJECTIVE: This study aimed to establish a highly sensitive and rapid single-tube, two-stage, multiplex recombinase-aided qPCR (mRAP) assay to specifically detect the khe, blaKPC-2, and blaNDM-1 genes in Klebsiella pneumoniae. METHODS: mRAP was carried out in a qPCR instrument within 1 h. The analytical sensitivities of mRAP for khe, blaKPC-2, and blaNDM-1 genes were tested using recombinant plasmids and dilutions of reference strains. A total of 137 clinical isolates and 86 sputum samples were used to validate the clinical performance of mRAP. RESULTS: mRAP achieved the sensitivities of 10, 8, and 14 copies/reaction for khe, blaKPC-2, and blaNDM-1 genes, respectively, superior to qPCR. The Kappa value of qPCR and mRAP for detecting khe, blaKPC-2, and blaNDM-1 genes was 1, 0.855, and 1, respectively (p < 0.05). CONCLUSION: mRAP is a rapid and highly sensitive assay for potential clinical identification of khe, blaKPC-2, and blaNDM-1 genes in K. pneumoniae.

Human ß-defensins and their synthetic analogs: Natural defenders and prospective new drugs of oral health.

As a family of cationic host defense peptides, human ß-defensins (HBDs) are ubiquitous in the oral cavity and are mainly synthesized primarily by epithelial cells, serving as the primary barrier and aiming to prevent microbial invasion, inflammation, and disease while maintaining physiological homeostasis. In recent decades, there has been great interest in their biological functions, structure-activity relationships, mechanisms of action, and therapeutic potential in oral diseases. Meanwhile, researchers are dedicated to improving the properties of HBDs for clinical application. In this review, we first describe the classification, structural characteristics, functions, and mechanisms of HBDs. Next, we cover the role of HBDs and their synthetic analogs in oral diseases, including dental caries and pulp infections, periodontitis, peri-implantitis, fungal/viral infections and oral mucosal diseases, and oral squamous cell carcinoma. Finally, we discuss the limitations and challenges of clinical translation of HBDs and their synthetic analogs, including, but not limited to, stability, bioavailability, antimicrobial activity, resistance, and toxicity. Above all, this review summarizes the biological functions, mechanisms of action, and therapeutic potential of both natural HBDs and their synthetic analogs in oral diseases, as well as the challenges associated with clinical translation, thus providing substantial insights into the laboratory development and clinical application of HBDs in oral diseases.

A Spatiotemporal Controllable Biomimetic Skin for Accelerating Wound Repair.

Skin injury repair is a dynamic process involving a series of interactions over time and space. Linking human physiological processes with materials' changes poses a significant challenge. To match the wound healing process, a spatiotemporal controllable biomimetic skin is developed, which comprises a three-dimensional (3D) printed membrane as the epidermis, a cell-containing hydrogel as the dermis, and a cytokine-laden hydrogel as the hypodermis. In the initial stage of the biomimetic skin repair wound, the membrane frame aids wound closure through pre-tension, while cells proliferate within the hydrogel. Next, as the frame disintegrates over time, cells released from the hydrogel migrate along the residual membrane. Throughout the process, continuous cytokines release from the hypodermis hydrogel ensures comprehensive nourishment. The findings reveal that in the rat full-thickness skin defect model, the biomimetic skin demonstrated a wound closure rate eight times higher than the blank group, and double the collagen content, particularly in the early repair process. Consequently, it is reasonable to infer that this biomimetic skin holds promising potential to accelerate wound closure and repair. This biomimetic skin with mechanobiological effects and spatiotemporal regulation emerges as a promising option for tissue regeneration engineering.

A Cyclical Magneto-Responsive Massage Dressing for Wound Healing.

Tissue development is mediated by a combination of mechanical and biological signals. Currently, there are many reports on biological signals regulating repair. However, insufficient attention is paid to the process of mechanical regulation, especially the active mechanical regulation in vivo, which has not been realized. Herein, a novel dynamically regulated repair system for both in vitro and in vivo applications is developed, which utilizes magnetic nanoparticles as non-contact actuators to activate hydrogels. The magnetic hydrogel can be periodically activated and deformed to different amplitudes by a dynamic magnetic system. An in vitro skin model is used to explore the impact of different dynamic stimuli on cellular mechano-transduction signal activation and cell differentiation. Specifically, the effect of mechanical stimulation on the phenotypic transition of fibroblasts to myofibroblasts is investigated. Furthermore, in vivo results verify that dynamic massage can simulate and enhance the traction effect in skin defects, thereby accelerating the wound healing process by promoting re-epithelialization and mediating dermal contraction.

Delicately Reinforced k-Nearest Neighbor Classifier Combined With Expert Knowledge Applied to Abnormity Forecast in Electrolytic Cell.

As the profit and safety requirements become higher and higher, it is more and more necessary to realize an advanced intelligent analysis for abnormity forecast of the synthetical balance of material and energy (AF-SBME) on aluminum reduction cells (ARCs). Without loss of generality, AF-SBME belongs to classification problems. Its advanced intelligent analysis can be realized by high-performance data-driven classifiers. However, AF-SBME has some difficulties, including a high requirement for interpretability of data-driven classifiers, a small number, and decreasing-over-time correctness of training samples. In this article, based on a preferable data-driven classifier, which is called a reinforced k -nearest neighbor (R-KNN) classifier, a delicately R-KNN combined with expert knowledge (DR-KNN/CE) is proposed. It improves R-KNN in two ways, including using expert knowledge as external assistance and enhancing self-ability to mine and synthesize data knowledge. The related experiments on AF-SBME, where the relevant data are directly sampled from practical production, have demonstrated that the proposed DR-KNN/CE not only makes an effective improvement for R-KNN, but also has a more advanced performance compared with other existing high-performance data-driven classifiers.

Deep Learning-Based 3D Single-Cell Imaging Analysis Pipeline Enables Quantification of Cell-Cell Interaction Dynamics in the Tumor Microenvironment.

The three-dimensional (3D) tumor microenvironment (TME) comprises multiple interacting cell types that critically impact tumor pathology and therapeutic response. Efficient 3D imaging assays and analysis tools could facilitate profiling and quantifying distinctive cell-cell interaction dynamics in the TMEs of a wide spectrum of human cancers. Here, we developed a 3D live-cell imaging assay using confocal microscopy of patient-derived tumor organoids and a software tool, SiQ-3D (single-cell image quantifier for 3D), that optimizes deep learning (DL)-based 3D image segmentation, single-cell phenotype classification, and tracking to automatically acquire multidimensional dynamic data for different interacting cell types in the TME. An organoid model of tumor cells interacting with natural killer cells was used to demonstrate the effectiveness of the 3D imaging assay to reveal immuno-oncology dynamics as well as the accuracy and efficiency of SiQ-3D to extract quantitative data from large 3D image datasets. SiQ-3D is Python-based, publicly available, and customizable to analyze data from both in vitro and in vivo 3D imaging. The DL-based 3D imaging analysis pipeline can be employed to study not only tumor interaction dynamics with diverse cell types in the TME but also various cell-cell interactions involved in other tissue/organ physiology and pathology. SIGNIFICANCE: A 3D single-cell imaging pipeline that quantifies cancer cell interaction dynamics with other TME cell types using primary patient-derived samples can elucidate how cell-cell interactions impact tumor behavior and treatment responses.

EGFR Inhibition Overcomes Resistance to FGFR4 Inhibition and Potentiates FGFR4 Inhibitor Therapy in Hepatocellular Carcinoma.

Aberrant activation of the FGF19-FGFR4 signaling pathway plays an essential role in the tumorigenesis of hepatocellular carcinoma (HCC). As such, FGFR4 inhibition has emerged as a novel therapeutic option for the treatment of HCC and has shown preliminary efficacy in recent clinical trials for patients exhibiting aberrant FGF19 expression. Resistance to kinase inhibitors is common in oncology, presenting a major challenge in the clinical treatment process. Hence, we investigated the potential mechanisms mediating and causing resistance to FGFR4 inhibition in HCC. Upon the successful establishment of a battery of cellular models developing resistance to FGFR4 inhibitors, we have identified the activation of EGFR, MAPK, and AKT signaling as the primary mechanisms mediating the acquired resistance. Combination of inhibitors against EGFR or its downstream components restored sensitivity to FGFR4 inhibitors. In parental HCC cell lines, EGF treatment also resulted in resistance to FGFR4 inhibitors. This resistance was effectively reverted by inhibitors of the EGFR signaling pathway, suggesting that EGFR activation is a potential cause of intrinsic resistance. We further confirmed the above findings in vivo in mouse xenograft tumor models. Genomic analysis of patient samples from The Cancer Genome Atlas confirmed that a segment of patients with HCC harboring FGF19 overexpression indeed exhibited increased activation of EGFR signaling. These findings conclusively indicate that both induced and innate activation of EGFR could mediate resistance to FGFR4 inhibition, suggesting that dual blockade of EGFR and FGFR4 may be a promising future therapeutic strategy for the treatment of FGF19-FGFR4 altered HCC.

Hyaluronic acid-mediated collagen intrafibrillar mineralization and enhancement of dentin remineralization.

Non-collagenous proteins (NCPs) in the extracellular matrix (ECM) of bone and dentin are known to play a critical regulatory role in the induction of collagen fibril mineralization and are embedded in hyaluronic acid (HA), which acts as a water-retaining glycosaminoglycan and provides necessary biochemical and biomechanical cues. Our previous study demonstrated that HA could regulate the mineralization degree and mechanical properties of collagen fibrils, yet its kinetics dynamic mechanism on mineralization is under debate. Here, we further investigated the role of HA on collagen fibril mineralization and the possible mechanism. The HA modification can significantly promote intrafibrillar collagen mineralization by reducing the electronegativity of the collagen surface to enhance calcium ions (Ca2+) binding capacity to create a local higher supersaturation. In addition, the HA also provides additional nucleation sites and shortens the induction time of amorphous calcium phosphate (ACP)-mediated hydroxyapatite (HAP) crystallization, which benefits mineralization. The acceleration effect of HA on intrafibrillar collagen mineralization is also confirmed in collagen hydrogel and in vitro dentin remineralization. These findings offer a physicochemical view of the regulation effect of carbohydrate polymers in the body on biomineralization, the fine prospect for an ideal biomaterial to repair collagen-mineralized tissues.

An injectable photopolymerizable chitosan hydrogel doped anti-inflammatory peptide for long-lasting periodontal pocket delivery and periodontitis therapy.

Periodontitis is a common chronic inflammatory disease caused by plaque that leads to alveolar bone resorption and tooth loss. Inflammation control and achieving better tissue repair are the key to periodontitis treatment. In this study, human ß-Defensin 1 short motif Pep-B with inflammation inhibition and differentiation regulation properties, is firstly used in the treatment of periodontitis, and an injectable photopolymerizable Pep-B/chitosan methacryloyl composite hydrogel (CMSA/Pep-B) is constructed. We confirm that Pep-B improves inflammation, and restores osteogenic behavior and function of injured stem cells. CMSA/Pep-B has good injectability, fluidity and photopolymerizability, and can sustainably release Pep-B to maintain drug concentration in periodontal pockets. Furthermore, animal experiments showed that CMSA/Pep-B significantly ameliorated the inflammation of the periodontium and reduced the alveolar bone loss by decreasing inflammatory infiltration, osteoclast formation and collagen destruction. In conclusion, CMSA/Pep-B is envisaged to be a novel bioactive material or therapeutic drug for treating periodontitis.

Influence of Dentist-Related Risk Factors on Implant Survival Provided by Dentists Without Structured Training: Results of a Retrospective Cross-Sectional Study.

PURPOSE: To evaluate the cumulative survival rate (CSR) of implants placed by Chinese dentists who lacked structured training and to identify the dentist-related risk factors associated with implant failure. MATERIALS AND METHODS: Data from 2,036 patients who underwent implant-supported restoration at a university-affiliated stomatology hospital were collected. CSR was regarded as the dependent variable. Patient-related characteristics (age, sex, insertion site, and surgical complexity) and dentist-related factors (experience, number of implant brands used, education level, sex, and specialty) were collected as independent variables. The chi-square test was used to identify dentist-related factors for implant failure after addressing patient-related potential confounders using propensity score matching (PSM). Dentist- and patient-related risk factors were further analyzed using multivariable logistic regression within the subgroups. RESULTS: The CSRs were 98.48% for patients (with single or multiple implants) and 98.86% for implants after 48 to 60 months of observation. Dentists with < 5 years of experience and specialists in implant dentistry were significantly associated with implant failure after addressing potential patient-related confounders. Within the group of dentists with < 5 years of experience, complicated cases were the major risk factor. For the group of specialists in implant dentistry, < 5 years of experience and male patient were the major risk factors. CONCLUSION: New dentists (< 5 years of experience) and specialists in implant dentistry are considered to be dentist-related risk factors for implant failure. This confirms that a learning curve exists for new specialists to reach the level of proficiency and expertise. Int J Oral Maxillofac Implants 2023;38:553-561. doi: 10.11607/jomi.9969.

A mitotic NADPH upsurge promotes chromosome segregation and tumour progression in aneuploid cancer cells.

Redox metabolites have been observed to fluctuate through the cell cycle in cancer cells, but the functional impacts of such metabolic oscillations remain unknown. Here, we uncover a mitosis-specific nicotinamide adenine dinucleotide phosphate (NADPH) upsurge that is essential for tumour progression. Specifically, NADPH is produced by glucose 6-phosphate dehydrogenase (G6PD) upon mitotic entry, which neutralizes elevated reactive oxygen species (ROS) and prevents ROS-mediated inactivation of mitotic kinases and chromosome missegregation. Mitotic activation of G6PD depends on the phosphorylation of its co-chaperone protein BAG3 at threonine 285, which results in dissociation of inhibitory BAG3. Blocking BAG3T285 phosphorylation induces tumour suppression. A mitotic NADPH upsurge is present in aneuploid cancer cells with high levels of ROS, while nearly unobservable in near-diploid cancer cells. High BAG3T285 phosphorylation is associated with worse prognosis in a cohort of patients with microsatellite-stable colorectal cancer. Our study reveals that aneuploid cancer cells with high levels of ROS depend on a G6PD-mediated NADPH upsurge in mitosis to protect them from ROS-induced chromosome missegregation.

Intrafibrillar Mineralization and Immunomodulatory for Synergetic Enhancement of Bone Regeneration via Calcium Phosphate Nanocluster Scaffold.

Inspired by the bionic mineralization theory, organic-inorganic composites with hydroxyapatite nanorods orderly arranged along collagen fibrils have attracted extensive attention. Planted with an ideal bone scaffold will contribute greatly to the osteogenic microenvironment; however, it remains challenging to develop a biomimetic scaffold with the ability to promote intrafibrillar mineralization and simultaneous regulation of immune microenvironment in situ. To overcome these challenges, a scaffold containing ultra-small particle size calcium phosphate nanocluster (UsCCP) is prepared, which can enhance bone regeneration through the synergetic effect of intrafibrillar mineralization and immunomodulatory. By efficient infiltration into collagen fibrils, the UsCCP released from the scaffold achieves intrafibrillar mineralization. It also promotes the M2-type polarization of macrophages, leading to an immune microenvironment with both osteogenic and angiogenic potential. The results confirm that the UsCCP scaffold has both intrafibrillar mineralization and immunomodulatory effects, making it a promising candidate for bone regeneration.

Levocarnitine regulates the growth of angiotensin II-induced myocardial fibrosis cells via TIMP-1.

This study aimed to explore the effects of tissue inhibitor of metalloproteinases-1 (TIMP-1) on levocarnitine (LC)-mediated regulation of angiotensin II (AngII)-induced myocardial fibrosis (MF) and its underlying mechanisms. H9C2 cells were treated with AngII for 24 h to induce fibrosis. The cells were then treated with LC or transfected with TIMP-1-OE plasmid/si­TIMP-1. Cell apoptosis, viability, migration, and related gene expression were analyzed. AngII treatment significantly upregulated Axl, α-SMA, and MMP3 expression (P < 0.05) and downregulated STAT4 and TIMP1 expression (P < 0.05) relative to the control levels. After transfection, cells with TIMP-1 overexpression/knockdown were successfully established. Compared with that of the control, AngII significantly inhibited cell viability and cell migration while promoting cell apoptosis (P < 0.05). LC and TIMP-1-OE transfection further suppressed cell viability and migration induced by Ang II and upregulated apoptosis, whereas si-TIMP-1 had the opposite effect. Furthermore, LC and TIMP-1-OE transfection downregulated Axl, AT1R, α-SMA, collagen III, Bcl-2, and MMP3 expression caused by AngII and upregulated caspase 3, p53, and STAT4 expression, whereas si-TIMP-1 had the opposite effect. TIMP-1 is therefore a potential therapeutic target for delaying MF progression.

Cytotoxic and chemotactic dynamics of NK cells quantified by live-cell imaging.

Natural Killer (NK) cells detect and eliminate virus-infected cells and cancer cells, and are crucial players of the human immune defense system. Although the relevant molecular machineries involved in NK cell activation and NK-target cell interactions are largely known, how their collective signaling modulates the dynamic behaviors of NK cells, e.g., motility and cytotoxicity, and the rate-limiting kinetics involved are still in need of comprehensive investigations. In traditional bulk killing assays, heterogeneity and kinetic details of individual NK-target cell interactions are masked, seriously limiting analysis of the underlying dynamic mechanisms. Here we present detailed protocols of a number of live-cell imaging assays using fluorescent protein reporters and/or a live-cell dye that enable the acquisition of quantitative kinetic data at the single cell level for elucidating the mechanism underlying the interaction dynamics of primary human NK cells and epithelial cancer cells. Moreover, we discuss how the imaging data can be analyzed either alone or in combination to quantify and determine the key dynamic steps/intermediates involved in specific NK cell activity, e.g., NK cell cytotoxic modes and their associated kinetics, and NK cell motility toward different cancer targets. These live-cell imaging assays can be easily adapted to analyze the rate-limiting kinetics and heterogeneity of other cell-cell interaction dynamics, e.g., in T cell function.

Aurora A-mediated pyruvate kinase M2 phosphorylation promotes biosynthesis with glycolytic metabolites and tumor cell cycle progression.

Cancer cells have distinctive demands for intermediates from glucose metabolism for biosynthesis and energy in different cell cycle phases. However, how cell cycle regulators and glycolytic enzymes coordinate to orchestrate the essential metabolic processes are still poorly characterized. Here, we report a novel interaction between the mitotic kinase, Aurora A, and the glycolytic enzyme, pyruvate kinase M2 (PKM2), in the interphase of the cell cycle. We found Aurora A-mediated phosphorylation of PKM2 at threonine 45. This phosphorylation significantly attenuated PKM2 enzymatic activity by reducing its tetramerization and also promoted glycolytic flux and the branching anabolic pathways. Replacing the endogenous PKM2 with a nonphosphorylated PKM2 T45A mutant inhibited glycolysis, glycolytic branching pathways, and tumor growth in both in vitro and in vivo models. Together, our study revealed a new protumor function of Aurora A through modulating a rate-limiting glycolytic enzyme, PKM2, mainly during the S phase of the cell cycle. Our findings also showed that although both Aurora A and Aurora B kinase phosphorylate PKM2 at the same residue, the spatial and temporal regulations of the specific kinase and PKM2 interaction are context dependent, indicating intricate interconnectivity between cell cycle and glycolytic regulators.

Downregulation of TCF1 in HIV Infection Impairs T-cell Proliferative Capacity by Disrupting Mitochondrial Function.

Background: Despite the benefits of antiretroviral therapy (ART) for people with HIV, T-cell dysfunction cannot be fully restored. Metabolic dysregulation is associated with dysfunction of HIV-1-specific T-cells. Exploration of the factors regulating metabolic fitness can help reverse T-cell dysfunction and provide new insights into the underlying mechanism. Methods: In this study, HIV-infected individuals and HIV-negative control individuals (NCs) were enrolled. T-cell factor (TCF)1 expression in cells was determined by quantitative reverse-transcriptase polymerase chain reaction and flow cytometry. Relevant microarray data from the GEO database were analyzed to explore the underlying mechanism. The effects of TCF1 on T-cell function and metabolic function were assessed in vitro. Results: TCF7 mRNA expression in peripheral blood mononuclear cells was downregulated in rapid progressors compared with long-term non-progressors individuals and NCs. TCF1 expression on CD4+ and CD8+ T-cells was downregulated in treatment-naïve HIV-infected individuals compared with NCs. Interleukin (IL)2 production and proliferative capacity were impaired in TCF1 knockdown T-cells. Moreover, glycolytic capacity and mitochondrial respiratory function were decreased in TCF1 knockdown T-cells, and depolarized mitochondria were increased in TCF1 knockdown T-cells. Conclusion: Downregulation of TCF1 in HIV infection impairs T-cell proliferative capacity by disrupting mitochondrial function. These findings highlight the metabolic regulation as a pivotal mechanism of TCF1 in the regulation of T-cell dysfunction.

Defect Passivation by a Multifunctional Phosphate Additive toward Improvements of Efficiency and Stability of Perovskite Solar Cells.

The quality of perovskite films plays a crucial role in the performance of the corresponding devices. However, the commonly employed perovskite polycrystalline films often contain a high density of defects created during film production and cell operation, including unsaturated coordinated Pb2+ and Pb0, which can act as nonradiative recombination centers, thus reducing open-circuit voltage. Effectively eliminating both kinds of defects is an important subject of research to improve the power conversion efficiency (PCE). Here, we employ hydrogen octylphosphonate potassium (KHOP) as a multifunctional additive to passivate defects. The molecule is introduced into perovskite precursor solution to regulate the perovskite film growth process by coordinating with Pb, which can not only passivate the Pb2+ defect but also effectively inhibit the production of Pb0; at the same time, the presence of K+ reduces device hysteresis by inhibiting I- migration and finally realizes double passivation of Pb2+ and I--based defects. Moreover, the moderate hydrophobic alkyl chain in the molecule improves the moisture stability. Ultimately, the optimal efficiency can reach 22.21%.

Ipriflavone suppresses NLRP3 inflammasome activation in host response to biomaterials and promotes early bone healing.

AIM: Emerging studies have shown that immune response to biomaterial implants plays a central role in bone healing. Ipriflavone is clinically used for osteoporosis. However, the mechanism of ipriflavone in immune response to implants in early stages of osseointegration remains unclear. In this study, we aimed to investigate the potential role of ipriflavone in early bone healing process and uncover the underlying mechanism. MATERIALS AND METHODS: We carried out histological examination as well as analysis of proinflammatory cytokines and NLRP3 inflammasome activation in a tibial implantation mouse model with intra-peritoneal injection of ipriflavone. In addition, we explored the mechanism of ipriflavone in the regulation of NLRP3 inflammasome activation in macrophages. RESULTS: In vivo, ipriflavone ameliorated host inflammatory response related to NLRP3 inflammasome activation at implantation sites, characterized by reductions of inflammatory cell infiltration and proinflammatory cytokine interleukin-1ß levels. Ipriflavone treatment also showed beneficial effects on early osseointegration. Further investigations of the molecular mechanism showed that the suppression of NLRP3 inflammasome acts upstream of NLRP3 oligomerization through abrogating the production of reactive oxygen species. CONCLUSIONS: These results revealed an anti-inflammatory role of ipriflavone in NLRP3 inflammasome activation through improving mitochondrial function. This study provides a new strategy for the development of immune-regulated biomaterials and treatment options for NLRP3-related diseases.