Palmitoylation licenses RIPK1 kinase activity and cytotoxicity in the TNF pathway.

Tumor necrosis factor (TNF)-induced receptor-interacting serine/threonine protein kinase 1 (RIPK1)-mediated cell death, including apoptosis and necroptosis, is increasingly recognized as a major driver of inflammatory diseases. Cell death checkpoints normally suppress RIPK1 kinase to safeguard the organism from its detrimental consequences. However, the mechanisms licensing RIPK1 kinase activity when a protective checkpoint is disabled remain unclear. Here, we identified S-palmitoylation as a licensing modification for RIPK1 kinase. TNF induces RIPK1 palmitoylation, mediated by DHHC5 and dependent on K63-linked ubiquitination of RIPK1, which enhances RIPK1 kinase activity by promoting the homo-interaction of its kinase domain and promotes cell death upon cell death checkpoint blockade. Furthermore, DHHC5 is amplified by fatty acid in the livers of mice with metabolic dysfunction-associated steatohepatitis, contributing to increased RIPK1 cytotoxicity observed in this condition. Our findings reveal that ubiquitination-dependent palmitoylation licenses RIPK1 kinase activity to induce downstream cell death signaling and suggest RIPK1 palmitoylation as a feasible target for inflammatory diseases.

Suppression of Enhancer Overactivation by a RACK7-Histone Demethylase Complex.

Regulation of enhancer activity is important for controlling gene expression programs. Here, we report that a biochemical complex containing a potential chromatin reader, RACK7, and the histone lysine 4 tri-methyl (H3K4me3)-specific demethylase KDM5C occupies many active enhancers, including almost all super-enhancers. Loss of RACK7 or KDM5C results in overactivation of enhancers, characterized by the deposition of H3K4me3 and H3K27Ac, together with increased transcription of eRNAs and nearby genes. Furthermore, loss of RACK7 or KDM5C leads to de-repression of S100A oncogenes and various cancer-related phenotypes. Our findings reveal a RACK7/KDM5C-regulated, dynamic interchange between histone H3K4me1 and H3K4me3 at active enhancers, representing an additional layer of regulation of enhancer activity. We propose that RACK7/KDM5C functions as an enhancer "brake" to ensure appropriate enhancer activity, which, when compromised, could contribute to tumorigenesis.

MYC Controls the Epstein-Barr Virus Lytic Switch.

Epstein-Barr virus (EBV) is associated with multiple human malignancies. To evade immune detection, EBV switches between latent and lytic programs. How viral latency is maintained in tumors or in memory B cells, the reservoir for lifelong EBV infection, remains incompletely understood. To gain insights, we performed a human genome-wide CRISPR/Cas9 screen in Burkitt lymphoma B cells. Our analyses identified a network of host factors that repress lytic reactivation, centered on the transcription factor MYC, including cohesins, FACT, STAGA, and Mediator. Depletion of MYC or factors important for MYC expression reactivated the lytic cycle, including in Burkitt xenografts. MYC bound the EBV genome origin of lytic replication and suppressed its looping to the lytic cycle initiator BZLF1 promoter. Notably, MYC abundance decreases with plasma cell differentiation, a key lytic reactivation trigger. Our results suggest that EBV senses MYC abundance as a readout of B cell state and highlights Burkitt latency reversal therapeutic targets.

G-quadruplexes on chromosomal DNA negatively regulates topoisomerase 1 activity.

Human DNA topoisomerase 1 (Top1) is a crucial enzyme responsible for alleviating torsional stress on DNA during transcription and replication, thereby maintaining genome stability. Previous researches had found that non-working Top1 interacted extensively with chromosomal DNA in human cells. However, the reason for its retention on chromosomal DNA remained unclear. In this study, we discovered a close association between Top1 and chromosomal DNA, specifically linked to the presence of G-quadruplex (G4) structures. G4 structures, formed during transcription, trap Top1 and hinder its ability to relax neighboring DNAs. Disruption of the Top1-G4 interaction using G4 ligand relieved the inhibitory effect of G4 on Top1 activity, resulting in a further reduction of R-loop levels in cells. Additionally, the activation of Top1 through the use of a G4 ligand enhanced the toxicity of Top1 inhibitors towards cancer cells. Our study uncovers a negative regulation mechanism of human Top1 and highlights a novel pathway for activating Top1.

Assembly of CpcL-phycobilisomes.

CpcL-phycobilisomes (CpcL-PBSs) are a reduced type of phycobilisome (PBS) found in several cyanobacteria. They lack the traditional PBS terminal energy emitters, but still show the characteristic red-shifted fluorescence at ~670 nm. We established a method of assembling in vitro a rod-membrane linker protein, CpcL, with phycocyanin, generating complexes with the red-shifted spectral features of CpcL-PBSs. The red-shift arises from the interaction of a conserved key glutamine, Q57 of CpcL in Synechocystis sp. PCC 6803, with a single phycocyanobilin chromophore of trimeric phycocyanin at one of the three ß82-sites. This chromophore is the terminal energy acceptor of CpcL-PBSs and donor to the photosystem(s). This mechanism also operates in PBSs from Acaryochloris marina MBIC11017. We then generated multichromic complexes harvesting light over nearly the complete visible range via the replacement of phycocyanobilin chromophores at sites α84 and ß153 of phycocyanins by phycoerythrobilin and/or phycourobilin. The results demonstrate the rational design of biliprotein-based light-harvesting elements by engineering CpcL and phycocyanins, which broadens the light-harvesting range and accordingly improves the light-harvesting capacity and may be potentially applied in solar energy harvesting.

Metabolomics and transcriptomics analysis revealed the response mechanism of alfalfa to combined cold and saline-alkali stress.

Cold and saline-alkali stress are frequently encountered by plants, and they often occur simultaneously in saline-alkali soils at mid to high latitudes, constraining forage crop distribution and production. However, the mechanisms by which forage crops respond to the combination of cold and saline-alkali stress remain unknown. Alfalfa (Medicago sativa L.) is one of the most essential forage grasses in the world. In this study, we analyzed the complex response mechanisms of two alfalfa species (Zhaodong [ZD] and Blue Moon [BM]) to combined cold and saline-alkali stress using multi-omics. The results revealed that ZD had a greater ability to tolerate combined stress than BM. The tricarboxylic acid cycles of the two varieties responded positively to the combined stress, with ZD accumulating more sugars, amino acids, and jasmonic acid. The gene expression and flavonoid content of the flavonoid biosynthesis pathway were significantly different between the two varieties. Weighted gene co-expression network analysis and co-expression network analysis based on RNA-Seq data suggested that the MsMYB12 gene may respond to combined stress by regulating the flavonoid biosynthesis pathway. MsMYB12 can directly bind to the promoter of MsFLS13 and promote its expression. Moreover, MsFLS13 overexpression can enhance flavonol accumulation and antioxidant capacity, which can improve combined stress tolerance. These findings provide new insights into improving alfalfa resistance to combined cold and saline-alkali stress, showing that flavonoids are essential for plant resistance to combined stresses, and provide theoretical guidance for future breeding programs.

MsMIOX2, encoding a MsbZIP53-activated myo-inositol oxygenase, enhances saline-alkali stress tolerance by regulating cell wall pectin and hemicellulose biosynthesis in alfalfa.

Alfalfa is one of the most widely cultivated forage crops worldwide. However, soil salinization restricts alfalfa growth and development and affects global productivity. The plant cell wall is the first barrier against various stresses. Therefore, elucidating the alterations in cell wall architecture is crucial for stress adaptation. This study aimed to clarify the impact of myo-inositol oxygenase 2 (MsMIOX2) on cell wall pectin and hemicellulose biosynthesis under saline-alkali stress and identify the upstream transcription factors that govern MsMIOX2. MsMIOX2 activation induced cell wall pectin and hemicellulose accumulation under saline-alkali stress. The effects of MsMIOX2 in saline-alkali tolerance were investigated by characterizing its overexpression and RNA interference lines. MsMIOX2 overexpression positively regulated the antioxidant system and photosynthesis in alfalfa under saline-alkali stress. MsMIOX2 exhibited myo-inositol oxygenase activity, which increased polysaccharide contents, facilitated pectin and hemicellulose biosynthesis, and extended the cell wall thickness. However, MsMIOX2 RNA interference decreased cell wall thickness and alleviated alfalfa saline-alkali stress tolerance. In addition, MsbZIP53 was identified as an upstream transcriptional MsMIOX2 regulator by yeast one-hybrid, electrophoretic mobility shift assay, dual-luciferase, and beta-glucuronidase assays. MsbZIP53 overexpression increased MsMIOX2 expression, elevated MIOX activity, reinforced the antioxidant system and photosynthesis, and increased saline-alkali stress tolerance in alfalfa. In conclusion, this study presents a novel perspective for elucidating the molecular mechanisms of saline-alkali stress tolerance in alfalfa and emphasizes the potential use of MsMIOX2 in alfalfa breeding.

Bi-allelic human TEKT3 mutations cause male infertility with oligoasthenoteratozoospermia owing to acrosomal hypoplasia and reduced progressive motility.

Oligoasthenoteratozoospermia (OAT) can result in male infertility owing to reduced sperm motility and abnormal spermatozoan morphology. The Tektins are a family of highly conserved filamentous proteins expressed in the axoneme and associated structures in many different metazoan species. Earlier studies on mice identified Tektin3 (Tekt3) as a testis-enriched gene, and knockout of Tekt3 resulted in asthenozoospermia in the mice. Here, whole-exome sequencing of 100 males with asthenozoospermia from unrelated families was performed, followed by Sanger sequencing, leading to the identification of TEKT3 as a candidate gene in two of these patients and their associated family members. In total, three mutations in the TEKT3 gene were identified in both these patients, including one homozygous deletion-insertion mutation (c.543_547delinsTTGAT: p.Glu182*) and one compound heterozygous mutation (c.[548G > A]; [752A > C], p.[Arg183Gln]; [Gln251Pro]). Both of these mutations resulted in the complete loss of TEKT3 expression. The patients were both found to produce sperm that, although those showed no apparent defects in the flagellar structure, had reduced progressive motility. In contrast to mice, most sperm from these two patients exhibited acrosomal hypoplasia, although this did not prevent the use of the sperm for in vitro fertilization through an ICSI approach. TEKT3 was found to bind to other TEKT proteins, suggesting that these proteins form a complex within human spermatozoa. Overall, these results suggest that a loss of TEKT3 function can contribute to OAT incidence in humans. TEKT3 deficiencies can reduce sperm motility and contribute to severe acrosomal hypoplasia in spermatozoa, compromising their normal function.

Induction-concurrent chemoradiotherapy with or without sintilimab in patients with locoregionally advanced nasopharyngeal carcinoma in China (CONTINUUM): a multicentre, open-label, parallel-group, randomised, controlled, phase 3 trial.

BACKGROUND: Anti-PD-1 therapy and chemotherapy is a recommended first-line treatment for recurrent or metastatic nasopharyngeal carcinoma, but the role of PD-1 blockade remains unknown in patients with locoregionally advanced nasopharyngeal carcinoma. We assessed the addition of sintilimab, a PD-1 inhibitor, to standard chemoradiotherapy in this patient population. METHODS: This multicentre, open-label, parallel-group, randomised, controlled, phase 3 trial was conducted at nine hospitals in China. Adults aged 18-65 years with newly diagnosed high-risk non-metastatic stage III-IVa locoregionally advanced nasopharyngeal carcinoma (excluding T3-4N0 and T3N1) were eligible. Patients were randomly assigned (1:1) using blocks of four to receive gemcitabine and cisplatin induction chemotherapy followed by concurrent cisplatin radiotherapy (standard therapy group) or standard therapy with 200 mg sintilimab intravenously once every 3 weeks for 12 cycles (comprising three induction, three concurrent, and six adjuvant cycles to radiotherapy; sintilimab group). The primary endpoint was event-free survival from randomisation to disease recurrence (locoregional or distant) or death from any cause in the intention-to-treat population. Secondary endpoints included adverse events. This trial is registered with ClinicalTrials.gov (NCT03700476) and is now completed; follow-up is ongoing. FINDINGS: Between Dec 21, 2018, and March 31, 2020, 425 patients were enrolled and randomly assigned to the sintilimab (n=210) or standard therapy groups (n=215). At median follow-up of 41·9 months (IQR 38·0-44·8; 389 alive at primary data cutoff [Feb 28, 2023] and 366 [94%] had at least 36 months of follow-up), event-free survival was higher in the sintilimab group compared with the standard therapy group (36-month rates 86% [95% CI 81-90] vs 76% [70-81]; stratified hazard ratio 0·59 [0·38-0·92]; p=0·019). Grade 3-4 adverse events occurred in 155 (74%) in the sintilimab group versus 140 (65%) in the standard therapy group, with the most common being stomatitis (68 [33%] vs 64 [30%]), leukopenia (54 [26%] vs 48 [22%]), and neutropenia (50 [24%] vs 46 [21%]). Two (1%) patients died in the sintilimab group (both considered to be immune-related) and one (<1%) in the standard therapy group. Grade 3-4 immune-related adverse events occurred in 20 (10%) patients in the sintilimab group. INTERPRETATION: Addition of sintilimab to chemoradiotherapy improved event-free survival, albeit with higher but manageable adverse events. Longer follow-up is necessary to determine whether this regimen can be considered as the standard of care for patients with high-risk locoregionally advanced nasopharyngeal carcinoma. FUNDING: National Natural Science Foundation of China, Key-Area Research and Development Program of Guangdong Province, Natural Science Foundation of Guangdong Province, Overseas Expertise Introduction Project for Discipline Innovation, Guangzhou Municipal Health Commission, and Cancer Innovative Research Program of Sun Yat-sen University Cancer Center. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.

Association of novel DNAH11 variants with asthenoteratozoospermia lead to male infertility.

BACKGROUND: Bi-allelic variants in DNAH11 have been identified as causative factors in Primary Ciliary Dyskinesia, leading to abnormal respiratory cilia. Nonetheless, the specific impact of these variants on human sperm flagellar and their involvement in male infertility remain largely unknown. METHODS: A collaborative effort involving two Chinese reproductive centers conducted a study with 975 unrelated infertile men. Whole-exome sequencing was employed for variant screening, and Sanger sequencing confirmed the identified variants. Morphological and ultrastructural analyses of sperm were conducted using Scanning Electron Microscopy and Transmission Electron Microscopy. Western Blot Analysis and Immunofluorescence Analysis were utilized to assess protein levels and localization. ICSI was performed to evaluate its efficacy in achieving favorable pregnancy outcomes for individuals with DNAH11 variants. RESULTS: In this study, we identified seven novel variants in the DNAH11 gene in four asthenoteratozoospermia subjects. These variants led the absence of DNAH11 proteins and ultrastructure defects in sperm flagella, particularly affecting the outer dynein arms (ODAs) and adjacent structures. The levels of ODA protein DNAI2 and axoneme related proteins were down regulated, instead of inner dynein arms (IDA) proteins DNAH1 and DNAH6. Two out of four individuals with DNAH11 variants achieved clinical pregnancies through ICSI. The findings confirm the association between male infertility and bi-allelic deleterious variants in DNAH11, resulting in the aberrant assembly of sperm flagella and contributing to asthenoteratozoospermia. Importantly, ICSI emerges as an effective intervention for overcoming reproductive challenges caused by DNAH11 gene variants.

Bi-allelic variants in chromatoid body protein TDRD6 cause spermiogenesis defects and severe oligoasthenoteratozoospermia in humans.

BACKGROUND: The association between the TDRD6 variants and human infertility remains unclear, as only one homozygous missense variant of TDRD6 was found to be associated with oligoasthenoteratozoospermia (OAT). METHODS: Whole-exome sequencing and Sanger sequencing were employed to identify potential pathogenic variants of TDRD6 in infertile men. Histology, immunofluorescence, immunoblotting and ultrastructural analyses were conducted to clarify the structural and functional abnormalities of sperm in mutated patients. Tdrd6-knockout mice were generated using the CRISPR-Cas9 system. Total RNA-seq and single-cell RNA-seq (scRNA-seq) analyses were used to elucidate the underlying molecular mechanisms, followed by validation through quantitative RT-PCR and immunostaining. Intracytoplasmic sperm injection (ICSI) was also used to assess the efficacy of clinical treatment. RESULTS: Bi-allelic TDRD6 variants were identified in five unrelated Chinese individuals with OAT, including homozygous loss-of-function variants in two consanguineous families. Notably, besides reduced concentrations and impaired motility, a significant occurrence of acrosomal hypoplasia was detected in multiple spermatozoa among five patients. Using the Tdrd6-deficient mice, we further elucidate the pivotal role of TDRD6 in spermiogenesis and acrosome identified. In addition, the mislocalisation of crucial chromatoid body components DDX4 (MVH) and UPF1 was also observed in round spermatids from patients harbouring TDRD6 variants. ScRNA-seq analysis of germ cells from a patient with TDRD6 variants revealed that TDRD6 regulates mRNA metabolism processes involved in spermatid differentiation and cytoplasmic translation. CONCLUSION: Our findings strongly suggest that TDRD6 plays a conserved role in spermiogenesis and confirms the causal relationship between TDRD6 variants and human OAT. Additionally, this study highlights the unfavourable ICSI outcomes in individuals with bi-allelic TDRD6 variants, providing insights for potential clinical treatment strategies.

miR-309a is a regulator of ovarian development in the oriental fruit fly Bactrocera dorsalis.

Fecundity is arguably one of the most important life history traits, as it is closely tied to fitness. Most arthropods are recognized for their extreme reproductive capacity. For example, a single female of the oriental fruit fly Bactrocera dorsalis, a highly invasive species that is one of the most destructive agricultural pests worldwide, can lay more than 3000 eggs during its life span. The ovary is crucial for insect reproduction and its development requires further investigation at the molecular level. We report here that miR-309a is a regulator of ovarian development in B. dorsalis. Our bioinformatics and molecular studies have revealed that miR-309a binds the transcription factor pannier (GATA-binding factor A/pnr), and this activates yolk vitellogenin 2 (Vg 2) and vitellogenin receptor (VgR) advancing ovarian development. We further show that miR-309a is under the control of juvenile hormone (JH) and independent from 20-hydroxyecdysone. Thus, we identified a JH-controlled miR-309a/pnr axis that regulates Vg2 and VgR to control the ovarian development. This study has further enhanced our understanding of molecular mechanisms governing ovarian development and insect reproduction. It provides a background for identifying targets for controlling important Dipteran pests.

Tailoring Highly Branched Poly(ß-amino ester)s for Efficient and Organ-Selective mRNA Delivery.

Development of mRNA therapeutics necessitates targeted delivery technology, while the clinically advanced lipid nanoparticles face difficulty for extrahepatic delivery. Herein, we design highly branched poly(ß-amino ester)s (HPAEs) for efficacious organ-selective mRNA delivery through tailoring their chemical compositions and topological structures. Using an "A2+B3+C2" Michael addition platform, a combinatorial library of 219 HPAEs with varied backbone structures, terminal groups, and branching degrees are synthesized. The branched topological structures of HPAEs provide enhanced serum resistance and significantly higher mRNA expression in vivo. The terminal amine structures of HPAEs determine the organ-selectivity of mRNA delivery following systemic administration: morpholine facilitates liver targeting, ethylenediamine favors spleen delivery, while methylpentane enables mRNA delivery to the liver, spleen, and lungs simultaneously. This study represents a comprehensive exploration of the structure-activity relationship governing both the efficiency and organ-selectivity of mRNA delivery by HPAEs, suggesting promising candidates for treating various organ-related diseases.

High Performance Nacre Fibers by Engineering Interfacial Entanglement.

Biological materials exhibit fascinating mechanical properties for intricate interactions at multiple interfaces to combine superb toughness with wondrous strength and stiffness. Recently, strong interlayer entanglement has emerged to replicate the powerful dissipation of natural proteins and alleviate the conflict between strength and toughness. However, designing intricate interactions in a strong entanglement network needs to be further explored. Here, we modulate interlayer entanglement by introducing multiple interactions, including hydrogen and ionic bonding, and achieve ultrahigh mechanical performance of graphene-based nacre fibers. Two essential modulating trends are directed. One is modulating dynamic hydrogen bonding to improve the strength and toughness up to 1.58 GPa and 52 MJ/m3, simultaneously. The other is tailoring ionic coordinating bonding to raise the strength and stiffness, reaching 2.3 and 253 GPa. Modulating various interactions within robust entanglement provides an effective approach to extend performance limits of bioinspired nacre and optimize multiscale interfaces in diverse composites.

Shared and distinct interactions of type 1 and type 2 Epstein-Barr Nuclear Antigen 2 with the human genome.

BACKGROUND: There are two major genetic types of Epstein-Barr Virus (EBV): type 1 (EBV-1) and type 2 (EBV-2). EBV functions by manipulating gene expression in host B cells, using virus-encoded gene regulatory proteins including Epstein-Barr Nuclear Antigen 2 (EBNA2). While type 1 EBNA2 is known to interact with human transcription factors (hTFs) such as RBPJ, EBF1, and SPI1 (PU.1), type 2 EBNA2 shares only ~ 50% amino acid identity with type 1 and thus may have distinct binding partners, human genome binding locations, and functions. RESULTS: In this study, we examined genome-wide EBNA2 binding in EBV-1 and EBV-2 transformed human B cells to identify shared and unique EBNA2 interactions with the human genome, revealing thousands of type-specific EBNA2 ChIP-seq peaks. Computational predictions based on hTF motifs and subsequent ChIP-seq experiments revealed that both type 1 and 2 EBNA2 co-occupy the genome with SPI1 and AP-1 (BATF and JUNB) hTFs. However, type 1 EBNA2 showed preferential co-occupancy with EBF1, and type 2 EBNA2 preferred RBPJ. These differences in hTF co-occupancy revealed possible mechanisms underlying type-specific gene expression of known EBNA2 human target genes: MYC (shared), CXCR7 (type 1 specific), and CD21 (type 2 specific). Both type 1 and 2 EBNA2 binding events were enriched at systemic lupus erythematosus (SLE) and multiple sclerosis (MS) risk loci, while primary biliary cholangitis (PBC) risk loci were specifically enriched for type 2 peaks. CONCLUSIONS: This study reveals extensive type-specific EBNA2 interactions with the human genome, possible differences in EBNA2 interaction partners, and a possible new role for type 2 EBNA2 in autoimmune disorders. Our results highlight the importance of considering EBV type in the control of human gene expression and disease-related investigations.

Roles of PD-L1 in human adipose-derived mesenchymal stem cells under inflammatory microenvironment.

Mesenchymal stem cells (MSCs) display unique homing and immunosuppression features which make them promising candidates for cell therapy in inflammatory disorders. It is known that C-X-C chemokine receptor type 4 (CXCR4, also known as CD184) is a critical receptor implicated in MSCs migration, and the protein programmed death ligand-1 (PD-L1) is involved in MSC's immunosuppression. However, it remains unclear how the molecular mechanisms regulate PD-L1 expression for migration and immunosuppression of MSCs under the inflammatory microenvironment. In this article, we used the human adipose-derived mesenchymal stem cells (hADMSCs) treated with lipopolysaccharide (LPS) as an in vitro inflammatory model to explore the roles of PD-L1 on the migration and immunosuppression of MSC. Our results demonstrate that in hADMSCs, LPS significantly increased PD-L1 expression, which mediated the migration of the LPS-treated hADMSCs via CXCR4. In addition, we found that the increased PD-L1 expression in the LPS-treated hADMSCs inhibited B cell proliferation and immunoglobulin G secretion through nuclear factor-κB. Our study suggests that the PD-L1 plays critical roles in the homing and immunosuppression of MSCs which are a promising cell therapy to treat inflammatory diseases.

Cyclic (Alkyl)(Amino)Carbene-Iminoboryl Compounds with Three Formal Oxidation States.

BN/CC isosterism is an effective strategy to build hybrid functional molecules with unique properties. In contrast to the alkynyl iminium salts derived from cyclic (alkyl)(amino)carbenes (CAACs) that feature only one reversible reduction wave, the isoelectronic cationic CAAC-iminoboryl adducts could be singly and doubly reduced smoothly. Both the resultant neutral radical and anionic azaborataallenes bear NBC-mixed allenic structures. The former radical has a high spin-density of 0.55e at CCAAC carbon, yet exhibits formal boron-centered radical reactivity. The latter azaborataallenes feature the nucleophilic CCAAC center and polar N(δ-)═B(δ+)═C(δ-) unit, and readily undergo nucleophilic substitution, isocyanide insertion, dipolar addition and cycloaddition reactions etc. The N-substituents have been shown to have a significant influence on the solid-state structure, thermal stability, and reactivity of azaborataallenes. This work showcases the allenic BN-unsaturated species as versatile building blocks in organic synthesis.

Senescent endothelial cells: a potential target for diabetic retinopathy.

Diabetic retinopathy (DR) is a diabetic complication that results in visual impairment and relevant retinal diseases. Current therapeutic strategies on DR primarily focus on antiangiogenic therapies, which particularly target vascular endothelial growth factor and its related signaling transduction. However, these therapies still have limitations due to the intricate pathogenesis of DR. Emerging studies have shown that premature senescence of endothelial cells (ECs) in a hyperglycemic environment is involved in the disease process of DR and plays multiple roles at different stages. Moreover, these surprising discoveries have driven the development of senotherapeutics and strategies targeting senescent endothelial cells (SECs), which present challenging but promising prospects in DR treatment. In this review, we focus on the inducers and mechanisms of EC senescence in the pathogenesis of DR and summarize the current research advances in the development of senotherapeutics and strategies that target SECs for DR treatment. Herein, we highlight the role played by key factors at different stages of EC senescence, which will be critical for facilitating the development of future innovative treatment strategies that target the different stages of senescence in DR.

Elucidating the biological characteristics and pathogenicity of the highly virulent G2a porcine epidemic diarrhea virus.

Since the large-scale outbreak of porcine epidemic diarrhoea (PED) in 2010, caused by the genotype 2 (G2) variant of the porcine epidemic diarrhoea virus (PEDV), pig farms in China, even those vaccinated with the G2b vaccine, have experienced infections from the G2a variant, leading to significant economic losses. This study successfully isolated the G2a strain DY2020 from positive small intestine contents (SICs) by blind passage on Vero cells for four generations. The SICs were taken from Daye, Hubei Province, China. The biological characteristics were identified by indirect immunofluorescence assay (IFA) and transmission electron microscopy (TEM). The growth kinetics of the strain on Vero cells were detected by TCID50, and the virus titre could reach 107.35 TCID50 ml-1 (SD: 5.07×106). The pathogenicity towards colostrum-deprived piglets was conducted by assessing faecal viral shedding, morphometric analysis of intestinal lesions, and immunohistochemical staining. The results showed that DY2020 was highly virulent to colostrum-deprived piglets, with severe watery diarrhoea and other clinical symptoms appeared at 6 h post-infection (h p.i.), and all died within 30 h. Pathological tissue examination results showed that the lesions mainly occurred in the intestines of piglets, causing pathological changes such as shortening of intestinal villi. In summary, the discovery of the G2a strain DY2020 in this study is of great significance for understanding Hubei PEDV and provides an important theoretical basis for the development of new efficient PEDV vaccines.

Enzyme Activity-Based Optical Probe: Imaging Aminopeptidases N in Vulnerable Plaque and Serum.

Cardiovascular disease, a chronic and progressive arterial wall disease, is increasingly recognized for its clinical significance. Aminopeptidases N (APN), crucial in the pathophysiological processes of vulnerable plaque, have been linked to endothelial dysfunction, oxidative stress, and plaque formation, thus highlighting their potential as biomarkers for disease progression. However, current detection methods for APN in body fluids and in vivo have limitations, including insufficient sensitivity and specificity, time delays, and the inability to directly reflect enzyme activity in plaques. To address these challenges, we developed an optical probe, HD-APN, for in vivo imaging of aminopeptidases, providing a potential implementation in cardiovascular disease. Our work demonstrated the applicability of HD-APN for specific monitoring of aminopeptidase levels in plaques and serum, shedding light on its potential for further research in cardiovascular disease.