o8G Site-Specifically Modified tRF-1-AspGTC: A Novel Therapeutic Target and Biomarker for Pulmonary Hypertension.

BACKGROUND: Hypoxia and oxidative stress contribute to the development of pulmonary hypertension (PH). tRNA-derived fragments play important roles in RNA interference and cell proliferation, but their epitranscriptional roles in PH development have not been investigated. We aimed to gain insight into the mechanistic contribution of oxidative stress-induced 8-oxoguanine in pulmonary vascular remodeling. METHODS: Through small RNA modification array analysis and quantitative polymerase chain reaction, a significant upregulation of the 8-oxoguanine -modified tRF-1-AspGTC was found in the lung tissues and the serum of patients with PH. RESULTS: This modification occurs at the position 5 of the tRF-1-AspGTC (5o8G tRF). Inhibition of the 5o8G tRF reversed hypoxia-induced proliferation and apoptosis resistance in pulmonary artery smooth muscle cells. Further investigation unveiled that the 5o8G tRF retargeted mRNA of WNT5A (Wingless-type MMTV integration site family, member 5A) and CASP3 (Caspase3) and inhibited their expression. Ultimately, BMPR2 (Bone morphogenetic protein receptor 2) -reactive oxygen species/5o8G tRF/WNT5A signaling pathway exacerbated the progression of PH. CONCLUSIONS: Our study highlights the role of site-specific 8-oxoguanine-modified tRF in promoting the development of PH. Our findings present a promising therapeutic avenue for managing PH and propose 5o8G tRF as a potential innovative marker for diagnosing this disease.

Nudix hydrolase WvNUDX24 is involved in borneol biosynthesis in Wurfbainia villosa.

Borneol, camphor, and bornyl acetate are highly promising monoterpenoids widely used in medicine, flavor, food, and chemical applications. Bornyl diphosphate (BPP) serves as a common precursor for the biosynthesis of these monoterpenoids. Although bornyl diphosphate synthase (BPPS) that catalyzes the cyclization of geranyl diphosphate (GPP) to BPP has been identified in multiple plants, the enzyme responsible for the hydrolysis of BPP to produce borneol has not been reported. Here, we conducted in vitro and in vivo functional characterization to identify the Nudix hydrolase WvNUDX24 from W. villosa, which specifically catalyzes the hydrolysis of BPP to generate bornyl phosphate (BP), and then BP forms borneol under the action of phosphatase. Subcellular localization experiments indicated that the hydrolysis of BPP likely occurs in the cytoplasm. Furthermore, site-directed mutagenesis experiments revealed that four critical residues (R84, S96, P98, and G99) for the hydrolysis activity of WvNUDX24. Additionally, the functional identification of phosphatidic acid phosphatase (PAP) demonstrated that WvPAP5 and WvPAP10 were able to hydrolyze geranylgeranyl diphosphate (GGPP) and farnesyl diphosphate (FPP) to generate geranylgeranyl phosphate (GGP) and farnesyl phosphate (FP), respectively, but could not hydrolyze BPP, GPP, and neryl diphosphate (NPP) to produce corresponding monophosphate products. These findings highlight the essential role of WvNUDX24 in the first step of BPP hydrolysis to produce borneol and provide genetic elements for the production of BPP-related terpenoids through plant metabolic engineering and synthetic biology.

ErbB3 Governs Endothelial Dysfunction in Hypoxia-Induced Pulmonary Hypertension.

BACKGROUND: Pulmonary hypertension, characterized by vascular remodeling, currently lacks curative therapeutic options. The dysfunction of pulmonary artery endothelial cells plays a pivotal role in the initiation and progression of pulmonary hypertension (PH). ErbB3 (human epidermal growth factor receptor 3), also recognized as HER3, is a member of the ErbB family of receptor tyrosine kinases. METHODS: Microarray, immunofluorescence, and Western blotting analyses were conducted to investigate the pathological role of ErbB3. Blood samples were collected for biomarker examination from healthy donors or patients with hypoxic PH. The pathological functions of ErbB3 were further validated in rodents subjected to chronic hypoxia- and Sugen-induced PH, with or without adeno-associated virus-mediated ErbB3 overexpression, systemic deletion, or endothelial cell-specific ErbB3 knockdown. Primary human pulmonary artery endothelial cells and pulmonary artery smooth muscle cells were used to elucidate the underlying mechanisms. RESULTS: ErbB3 exhibited significant upregulation in the serum, lungs, distal pulmonary arteries, and pulmonary artery endothelial cells isolated from patients with PH compared with those from healthy donors. ErbB3 overexpression stimulated hypoxia-induced endothelial cell proliferation, exacerbated pulmonary artery remodeling, elevated systolic pressure in the right ventricle, and promoted right ventricular hypertrophy in murine models of PH. Conversely, systemic deletion or endothelial cell-specific knockout of ErbB3 yielded opposite effects. Coimmunoprecipitation and proteomic analysis identified YB-1 (Y-box binding protein 1) as a downstream target of ErbB3. ErbB3 induced nuclear translocation of YB-1 and subsequently promoted hypoxia-inducible factor 1/2α transcription. A positive loop involving ErbB3-periostin-hypoxia-inducible factor 1/2α was identified to mediate the progressive development of this disease. MM-121, a human anti-ErbB3 monoclonal antibody, exhibited both preventive and therapeutic effects against hypoxia-induced PH. CONCLUSIONS: Our study reveals, for the first time, that ErbB3 serves as a novel biomarker and a promising target for the treatment of PH.

Cancer-Associated Endocrine Cells Participate in Pancreatic Carcinogenesis.

BACKGROUND & AIMS: The pancreas is composed of endocrine and exocrine parts, and its interlacing structure indicates potential interaction between endocrine and exocrine cells. Although the tumor microenvironment of pancreatic ductal adenocarcinoma (PDAC) has been well characterized, the role of pancreatic endocrine cells during carcinogenesis is relatively understudied. METHODS: The changes of endocrine cells in PDAC by single-cell transcriptome sequencing, spatial transcriptome sequencing, and multiplex immunohistochemistry were depicted. After that, the interaction between pancreatic carcinogenesis and endocrine changes was explored in orthotopic transplantation mice, KrasLSL-G12DPdx1-Cre mice, and KrasLSL-G12Dp53LoxPPdx1-CreER mice. Finally, we proved the mechanism of the interaction between endocrine and exocrine parts of the pancreas through islet isolation, co-culture in vitro and co-injection in vivo. RESULTS: Pancreatic endocrine cells displayed significantly different transcriptomic characteristics and increased interaction with exocrine part in PDAC. Specifically, among all of the changes, pancreatic polypeptide-positive cells showed a sharp increment accompanied by the progression of the cancer lesion, which might be derived from the transdifferentiation of α and ß cells. Interestingly, it was proved that PDAC cells were able to induce the transdifferentiation of pancreatic α cells and ß cells into glucagon-pancreatic polypeptide and insulin-pancreatic polypeptide double-positive cells, which further promoted carcinogenesis and development of PDAC in a paracrine-dependent manner and formed a reciprocal interaction. CONCLUSIONS: This study systematically maps the alteration of pancreatic endocrine cells in PDAC and elucidates the potential endocrine-exocrine interaction mechanisms during PDAC carcinogenesis. In addition, cancer-associated endocrine cells are defined and characterized, thereby further broadening the composition of PDAC microenvironment.

UPK3A+ umbrella cell damage mediated by TLR3-NR2F6 triggers programmed destruction of urothelium in Hunner-type interstitial cystitis/painful bladder syndrome.

Urothelial damage and barrier dysfunction emerge as the foremost mechanisms in Hunner-type interstitial cystitis/bladder pain syndrome (HIC). Although treatments aimed at urothelial regeneration and repair have been employed, their therapeutic effectiveness remains limited due to the inadequate understanding of specific cell types involved in damage and the lack of specific molecular targets within these mechanisms. Therefore, we harnessed single-cell RNA sequencing to elucidate the heterogeneity and developmental trajectory of urothelial cells within HIC bladders. Through reclustering, we identified eight distinct clusters of urothelial cells. There was a significant reduction in UPK3A+ umbrella cells and a simultaneous increase in progenitor-like pluripotent cells (PPCs) within the HIC bladder. Pseudotime analysis of the urothelial cells in the HIC bladder revealed that cells faced challenges in differentiating into UPK3A+ umbrella cells, while PPCs exhibited substantial proliferation to compensate for the loss of UPK3A+ umbrella cells. The urothelium in HIC remains unrepaired, despite the substantial proliferation of PPCs. Thus, we propose that inhibiting the pivotal signaling pathways responsible for the injury to UPK3A+ umbrella cells is paramount for restoring the urothelial barrier and alleviating lower urinary tract symptoms in HIC patients. Subsequently, we identified key molecular pathways (TLR3 and NR2F6) associated with the injury of UPK3A+ umbrella cells in HIC urothelium. Finally, we conducted in vitro and in vivo experiments to confirm the potential of the TLR3-NR2F6 axis as a promising therapeutic target for HIC. These findings hold the potential to inhibit urothelial injury, providing promising clues for early diagnosis and functional bladder self-repair strategies for HIC patients. © 2024 The Pathological Society of Great Britain and Ireland.

Impacts of pro-inflammatory cytokines variant on cardiometabolic profile and premature coronary artery disease: A systematic review and meta-analysis.

Interleukin-6 (IL-6), a pivotal pro-inflammatory cytokine, is closely linked to vascular wall thickening and atherosclerotic lesion. Since serum IL-6 levels are largely determined by the genetic variant in IL-6, this study was conducted to investigate whether the IL-6 variant impacts cardiometabolic profile and the risk of premature coronary artery disease (PCAD). PubMed, Cochrane Library, Central, Cumulative Index to Nursing and Allied Health Literature (CINAHL), and ClinicalTrials.gov were searched from May 13, 2022 to June 28, 2023. In total, 40 studies (26,543 individuals) were included for the analysis. The rs1800795 (a function variant in the IL-6 gene) C allele was linked to higher levels of low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), fasting plasma glucose (FPG), body mass index (BMI), and waist circumference (WC), and a lower levels of high-density lipoprotein cholesterol (HDL-C). However, no significant association was observed of rs1800795 with triglycerides (TG), systolic blood pressure (SBP), and diastolic blood pressure (DBP). Interestingly, a significant association was detected between rs1800795 and PCAD. Subgroup analyses indicted that the impacts of rs1800795 on cardiometabolic risk factors were significant in Caucasians but stronger in obese patients. In contrast, the impact of rs1800795 on PCAD was significant in brown race population. In summary, rs1800795 had a slight but significant impact on cardiometabolic risk factors and PCAD. IL-6 inhibition with ziltivekimab or canakinumab may benefit high-risk populations (e.g. brown race population, Caucasians, obese patients, etc.) with rs1800795 to prevent PCAD.

Unraveling the pH-Dependent Oxygen Reduction Performance on Single-Atom Catalysts: From Single- to Dual-Sabatier Optima.

Metal-nitrogen-carbon (M-N-C) single-atom catalysts (SACs) have emerged as a potential substitute for the costly platinum-group catalysts in oxygen reduction reaction (ORR). However, several critical aspects of M-N-C SACs in ORR remain poorly understood, including their pH-dependent activity, selectivity for 2- or 4-electron transfer pathways, and the identification of the rate-determining steps. Herein, by analyzing >100 M-N-C structures and >2000 sets of energetics, we unveil a pH-dependent evolution in ORR activity volcanos─from a single peak in alkaline media to a double peak in acids. We found that this pH-dependent behavior in M-N-C catalysts fundamentally stems from their moderate dipole moments and polarizability for O* and HOO* adsorbates, as well as unique scaling relations among ORR adsorbates. To validate our theoretical discovery, we synthesized a series of molecular M-N-C catalysts, each characterized by well-defined atomic coordination environments. Impressively, the experiments matched our theoretical predictions on kinetic current, Tafel slope, and turnover frequency in both acidic and alkaline environments. These new insights also refine the famous Sabatier principle by emphasizing the need to avoid an "acid trap" while designing M-N-C catalysts for ORR or any other pH-dependent electrochemical applications.

BPR3P0128, a non-nucleoside RNA-dependent RNA polymerase inhibitor, inhibits SARS-CoV-2 variants of concern and exerts synergistic antiviral activity in combination with remdesivir.

Viral RNA-dependent RNA polymerase (RdRp), a highly conserved molecule in RNA viruses, has recently emerged as a promising drug target for broad-acting inhibitors. Through a Vero E6-based anti-cytopathic effect assay, we found that BPR3P0128, which incorporates a quinoline core similar to hydroxychloroquine, outperformed the adenosine analog remdesivir in inhibiting RdRp activity (EC50 = 0.66 µM and 3 µM, respectively). BPR3P0128 demonstrated broad-spectrum activity against various severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern. When introduced after viral adsorption, BPR3P0128 significantly decreased SARS-CoV-2 replication; however, it did not affect the early entry stage, as evidenced by a time-of-drug-addition assay. This suggests that BPR3P0128's primary action takes place during viral replication. We also found that BPR3P0128 effectively reduced the expression of proinflammatory cytokines in human lung epithelial Calu-3 cells infected with SARS-CoV-2. Molecular docking analysis showed that BPR3P0128 targets the RdRp channel, inhibiting substrate entry, which implies it operates differently-but complementary-with remdesivir. Utilizing an optimized cell-based minigenome RdRp reporter assay, we confirmed that BPR3P0128 exhibited potent inhibitory activity. However, an enzyme-based RdRp assay employing purified recombinant nsp12/nsp7/nsp8 failed to corroborate this inhibitory activity. This suggests that BPR3P0128 may inhibit activity by targeting host-related RdRp-associated factors. Moreover, we discovered that a combination of BPR3P0128 and remdesivir had a synergistic effect-a result likely due to both drugs interacting with separate domains of the RdRp. This novel synergy between the two drugs reinforces the potential clinical value of the BPR3P0128-remdesivir combination in combating various SARS-CoV-2 variants of concern.

Structure and function of the pyridoxal 5'-phosphate-dependent (PLP) threonine deaminase IlvA1 from Pseudomonas aeruginosa PAO1.

IlvA1, a pyridoxal phosphate-dependent (PLP) enzyme, catalyzes the deamination of l-threonine and l-serine to yield 2-ketobutyric acid or pyruvate. To gain insights into the function of IlvA1, we determined its crystal structure from Pseudomonas aeruginosa to 2.3 Å. Density for a 2-ketobutyric acid product was identified in the active site and a putative allosteric site. Activity and substrate binding assays confirmed that IlvA1 utilizes l-threonine, l-serine, and L-allo-threonine as substrates. The enzymatic activity is regulated by the end products l-isoleucine and l-valine. Additionally, the efficiency of d-cycloserine and l-cycloserine inhibitors on IlvA1 enzymatic activity was examined. Notably, site-directed mutagenesis confirmed the active site residues and revealed that Gln165 enhances the enzyme activity, emphasizing its role in substrate access. This work provides crucial insights into the structure and mechanism of IlvA1 and serves as a starting point for further functional and mechanistic studies of the threonine deaminase in P. aeruginosa.

Maize/soybean intercropping increases nutrient uptake, crop yield and modifies soil physio-chemical characteristics and enzymatic activities in the subtropical humid region based in Southwest China.

Intercropping, a widely adopted agricultural practice worldwide, aims to increase crop yield, enhance plant nutrient uptake, and optimize the utilization of natural resources, contributing to sustainable farming practices on a global scale. However, the underlying changes in soil physio-chemical characteristics and enzymatic activities, which contribute to crop yield and nutrient uptake in the intercropping systems are largely unknown. Consequently, a two-year (2021-2022) field experiment was conducted on the maize/soybean intercropping practices with/without nitrogen (N) fertilization (i.e., N0; 0 N kg ha-1 and N1; 225 N kg ha-1 for maize and 100 N kg ha-1 for soybean ) to know whether such cropping system can improve the nutrients uptake and crop yields, soil physio-chemical characteristics, and soil enzymes, which ultimately results in enhanced crop yield. The results revealed that maize intercropping treatments (i.e., N0MI and N1MI) had higher crop yield, biomass dry matter, and 1000-grain weight of maize than mono-cropping treatments (i.e., N0MM, and N1MM). Nonetheless, these parameters were optimized in N1MI treatments in both years. For instance, N1MI produced the maximum grain yield (10,105 and 11,705 kg ha-1), biomass dry matter (13,893 and 14,093 kg ha-1), and 1000-grain weight (420 and 449 g) of maize in the year 2021 and 2022, respectively. Conversely, soybean intercropping treatments (i.e., N0SI and N1SI) reduced such yield parameters for soybean. Also, the land equivalent ratio (LER) and land equivalent ratio for N fertilization (LERN) values were always greater than 1, showing the intercropping system's benefits in terms of yield and improved resource usage. Moreover, maize intercropping treatments (i.e., N0MI and N1MI) and soybean intercropping treatments (i.e., N0SI and N1SI) significantly (p < 0.05) enhanced the nutrient uptake (i.e., N, P, K, Ca, Fe, and Zn) of maize and soybean, however, these nutrients uptakes were more prominent in N1MI and N1SI treatments of maize and soybean, respectively in both years (2021 and 2022) compared with their mono-cropping treatments. Similarly, maize-soybean intercropping treatments (i.e., N0MSI and N1MSI) significantly (p < 0.05) improved the soil-based N, P, K, NH4, NO3, and soil organic matter, but, reduced the soil pH. Such maize-soybean intercropping treatments also improved the soil enzymatic activities such as protease (PT), sucrose (SC), acid phosphatase (AP), urease (UE), and catalase (CT) activities. This indicates that maize-soybean intercropping could potentially contribute to higher and better crop yield, enhanced plant nutrient uptake, improved soil nutrient pool, physio-chemical characteristics, and related soil enzymatic activities. Thus, preferring intercropping to mono-cropping could be a preferable choice for ecologically viable agricultural development.

Low temperature exposure influences nitrogen metabolism resulting in decreased Cry1Ac insecticidal endotoxin content in cotton seeds.

BACKGROUND: Sudden temperature drops, resulting from extreme weather events, often occur during the boll-setting period of cotton in Xinjiang, China, causing decreased expression of Bacillus thuringiensis (Bt) insecticidal proteins in cotton bolls. The precise threshold temperatures and durations that lead to significant changes in Cry1Ac endotoxin levels under low temperatures remain unclear. To address this, we investigated the effects of different temperatures and stress durations on Cry1Ac endotoxin levels in cotton bolls. In 2020-2021, two Bt transgenic cotton varieties, conventional Sikang1 and hybrid Sikang3, were selected as experimental materials. Various low temperatures (ranging from 16 to 20 °C) with different durations (12 h, 24 h and 48 h) were applied during the peak boll-setting period. RESULTS: As the temperature decreased, the Cry1Ac endotoxin content in the boll shell, fiber, and seed exhibited a declining trend. Moreover, the threshold temperature which caused a significant reduction in Cry1Ac endotoxin content increased with the prolonged duration of low-temperature stress. Among the components of cotton bolls, seeds were most affected by low-temperature stress, with the threshold temperature for a significant reduction in Cry1Ac endotoxin content ranging from 17 °C to 19 °C. Correlation analysis indicated that low temperatures led to a decrease in protein synthesis capacity and an increase in degradation ability, resulting in reduced Cry1Ac endotoxin content. Pathway analysis revealed that both free amino acid and peptidase had significant negative effects on Cry1Ac endotoxin content. CONCLUSION: In summary, when the daily average temperature was ≤ 19 °C, implementing cultural practices to reduce free amino acid content and peptidase activity could serve as effective cold defense strategies for Bt cotton production.

Evaluation of the environmental factors influencing the quality of Astragalus membranaceus var. mongholicus based on HPLC and the Maxent model.

BACKGROUND: In recent years, global climate change in tandem with increased human activity has resulted in habitat degradation or the migration of rare medicinal plants, potentially impacting the quality of medicinal herbs. Astragalus membranaceus var. mongholicus is a valuable bulk medicinal material in Northwest China. As the demand for this medicinal herb continues to increase in both domestic and international markets, ensuring the sustainable development of high-quality Astragali Radix is important. In this study, the maximum entropy (Maxent) model was applied, thereby incorporating 136 distribution records, along with 39 environmental factors of A. membranaceus var. mongholicus, to assess the quality zonation and potential distribution of this species in China under climate change. RESULTS: The results showed that the elevation, annual mean temperature, precipitation of wettest month, solar radiation in June, and mean temperature of warmest quarter were the critical environmental factors influencing the accumulation of astragaloside IV and Astragalus polysaccharide in A. membranaceus var. mongholicus. Among the twelve main environmental variables, annual mean temperature, elevation, precipitation of the wettest month, and solar radiation in November were the four most important factors influencing the distribution of A. membranaceus var. mongholicus. In addition, ecological niche modelling revealed that highly suitable habitats were mainly located in central and western Gansu, eastern Qinghai, northern Shaanxi, southern Ningxia, central Inner Mongolia, central Shanxi, and northern Hebei. However, the future projections under climate change suggested a contraction of these suitable areas, shifting towards northeastern high-latitude and high-elevation mountains. CONCLUSIONS: The findings provide essential insights for developing adaptive strategies for A. membranaceus var. mongholicus cultivation in response to climate change and can inform future research on this species. By considering the identified environmental factors and the potential impacts of the predicted climate changes, we can visualize the regional distribution of high-quality Radix Astragali and develop conservation strategies to protect and restore its suitable habitats.

Impact of exercise on chemotherapy-induced peripheral neuropathy in survivors with post-treatment primary breast cancer.

PURPOSE: Chemotherapy-induced peripheral neuropathy (CIPN) is a debilitating side effect of neurotoxic chemotherapy. Exercise activates neuromuscular function and may improve CIPN. We examined the association between exercise and CIPN symptoms in breast cancer survivors. METHODS: In a retrospective cross-sectional study, we included patients completing a survey assessing exercise exposure and neuropathy symptoms in a tertiary cancer center survivorship clinic. We evaluated exercise duration and intensity using a standardized questionnaire quantified in metabolic equivalent tasks (MET-h/wk). We defined exercisers as patients meeting the National Physical Activity Guidelines' criteria. We used multivariable logistic regressions to examine the relationship between exercise and CIPN and if this differed as a function of chemotherapy regimen adjusting for age, gender, and race. RESULTS: We identified 5444 breast cancer survivors post-chemotherapy (median age 62 years (interquartile range [IQR]: 55, 71); median 4.7 years post-chemotherapy (IQR: 3.3, 7.6)) from 2017 to 2022. CIPN overall prevalence was 34% (95% confidence interval [CI]: 33%, 36%), 33% for non-taxane, and 37% for taxane-based chemotherapy. CIPN prevalence was 28% (95% CI: 26%, 30%) among exercisers and 38% (95% CI: 37%, 40%) among non-exercisers (difference 11%; 95% CI: 8%, 13%; p < 0.001). Compared to patients with low (<6 MET-h/wk) levels of exercise (42%), 11% fewer patients with moderate (6-20.24 MET-h/wk) to high (>20.25 MET-h/wk) levels of exercise reported CIPN. Exercise was associated with reduced prevalence of all CIPN symptoms regardless of chemotherapy type. CONCLUSION: CIPN may persist several years following chemotherapy among patients with breast cancer but is significantly reduced by exercise in a dose-dependent manner.

Platinum-Ruthenium Bimetallic Nanoparticle Catalysts Synthesized Via Direct Joule Heating for Methanol Fuel Cells.

Platinum-Ruthenium (PtRu) bimetallic nanoparticles are promising catalysts for methanol oxidation reaction (MOR) required by direct methanol fuel cells. However, existing catalyst synthesis methods have difficulty controlling their composition and structures. Here, a direct Joule heating method to yield highly active and stable PtRu catalysts for MOR is shown. The optimized Joule heating condition at 1000 °C over 50 microseconds produces uniform PtRu nanoparticles (6.32 wt.% Pt and 2.97 wt% Ru) with an average size of 2.0 ± 0.5 nanometers supported on carbon black substrates. They have a large electrochemically active surface area (ECSA) of 239 m2 g-1 and a high ECSA normalized specific activity of 0.295 mA cm-2. They demonstrate a peak mass activity of 705.9 mA mgPt -1 for MOR, 2.8 times that of commercial 20 wt.% platinum/carbon catalysts, and much superior to PtRu catalysts obtained by standard hydrothermal synthesis. Theoretical calculation results indicate that the superior catalytic activity can be attributed to modified Pt sites in PtRu nanoparticles, enabling strong methanol adsorption and weak carbon monoxide binding. Further, the PtRu catalyst demonstrates excellent stability in two-electrode methanol fuel cell tests with 85.3% current density retention and minimum Pt surface oxidation after 24 h.

Molecular and clinical characterization of two unrelated families with factor V deficiency, including a novel nonsense variant (p.Gln1532*).

BACKGROUND: Factor V (FV) is an essential cofactor in the coagulation cascade. The characterization of novel mutations is advantageous for the clinical management of FV-deficient patients. METHODS: Coagulation screening and thrombin generation assay were performed with the plate-poor plasma. All 25 exons of the F5 gene were amplified and sequenced. The ClustalX-2.1 software was applied to the multiple sequence alignment. The possible adverse effects of mutations were investigated with online bioinformatics software and protein modeling. RESULTS: Two unrelated families with FV deficiency were under investigation. Proband A was an 18-year-old youth with recurrent epistaxis. Proband B was a 29-year-old woman who did not present with any bleeding symptoms. Three heterozygous mutations (p.Gln1532*, p.Phe218Ser, and p.Asp2222Gly) were detected. Interestingly, they were compound heterozygotes and both contained the p.Asp2222Gly, a polymorphism. The thrombin generation assay showed that both patients had impaired ability of thrombin generation, and in particular, proband A was more severe. Conservation, pathogenicity and protein modeling studies all indicated that these three mutations could cause deleterious effects on the function and structure of FV. CONCLUSION: These three mutations are responsible for the FV-deficient in two pedigrees. Moreover, the nonsense variant p.Gln1532* is first reported in the world.

Progress of single-cell RNA sequencing combined with spatial transcriptomics in tumour microenvironment and treatment of pancreatic cancer.

In recent years, single-cell analyses have revealed the heterogeneity of the tumour microenvironment (TME) at the genomic, transcriptomic, and proteomic levels, further improving our understanding of the mechanisms of tumour development. Single-cell RNA sequencing (scRNA-seq) technology allow analysis of the transcriptome at the single-cell level and have unprecedented potential for exploration of the characteristics involved in tumour development and progression. These techniques allow analysis of transcript sequences at higher resolution, thereby increasing our understanding of the diversity of cells found in the tumour microenvironment and how these cells interact in complex tumour tissue. Although scRNA-seq has emerged as an important tool for studying the tumour microenvironment in recent years, it cannot be used to analyse spatial information for cells. In this regard, spatial transcriptomics (ST) approaches allow researchers to understand the functions of individual cells in complex multicellular organisms by understanding their physical location in tissue sections. In particular, in related research on tumour heterogeneity, ST is an excellent complementary approach to scRNA-seq, constituting a new method for further exploration of tumour heterogeneity, and this approach can also provide unprecedented insight into the development of treatments for pancreatic cancer (PC). In this review, based on the methods of scRNA-seq and ST analyses, research progress on the tumour microenvironment and treatment of pancreatic cancer is further explained.

Screening and experimental validation of diagnostic gene in ulcerative colitis with anti-TNF-α therapy.

In clinical practice, the diagnosis of ulcerative colitis (UC) mainly relies on a comprehensive analysis of a series of signs and symptoms of patients. The current biomarkers for diagnosis of UC and prognostic prediction of anti-TNF-α therapy are inaccurate. The present study aimed to perform an integrative analysis of gene expression profiles in patients with UC. A total of seven datasets from the GEO database that met our strict inclusion criteria were included. After identifying differentially expressed genes (DEGs) between UC patients and healthy individuals, the diagnostic and prognostic utility of the DEGs were then analyzed via least absolute shrinkage and selection operator and support-vector machine recursive feature elimination. Subgroup analyses of the treated and untreated groups, as well as the treatment-response group and non-response group, were also performed. Furthermore, the relationship between the expressions of UC-related genes and infiltration of immune cells in the course of treatment was also investigated. Immunohistochemical (IHC) assay was used to verify the gene expression in inflamed UC tissues. When considering all the applied methods, DUOX2, PI3, S100P, MMP7, and S100A8 had priority to be defined as the characteristic genes among DEGs. The area under curve (AUC) of the five genes, which were all consistently over-expressed, based on an external validation dataset, were all above 0.94 for UC diagnosis. Four of the five genes (DUOX2, PI3, MMP7, and S100A8) were down-regulated between treatment-responsive and nonresponsive patients. A significant difference was also observed concerning the infiltration of immune cells, including macrophage and neutrophil, between the two groups (treatment responsive and nonresponsive). The changes in the expression of DUOX2 and MMP7 based on the IHC assay were highly consistent with the results obtained in the current study. This confirmed the mild to moderate diagnostic and predictive value of DUOX2 and MMP7 in patients with UC. The conducted analyses showed that the expression profile of the five identified biomarkers accurately detects UC, whereas four of the five genes evidently predicted the response to anti-TNF-α therapy.

Dynamic m6A mRNA methylation reveals the involvement of AcALKBH10 in ripening-related quality regulation in kiwifruit.

Kiwifruit ripening is a complex and highly coordinated process that occurs in conjunction with the formation of fruit edible quality. The significance of epigenetic changes, particularly the impact of N6-methyladenosine (m6A) RNA modification on fruit ripening and quality formation, has been largely overlooked. We monitored m6A levels and gene expression changes in kiwifruit at four different stages using LC-MS/MS, MeRIP, RNA-seq, and validated the function of AcALKBH10 through heterologous transgenic expression in tomato. Notable m6A modifications occurred predominantly at the stop codons and the 3' UTRs and exhibited a gradual reduction in m6A levels during the fruit ripening process. Moreover, these m6A modifications in the aforementioned sites demonstrated a discernible inverse relationship with the levels of mRNA abundance throughout the ripening process, suggesting a repression effect of m6A modification in the modulation of kiwifruit ripening. We further demonstrated that AcALKBH10 rather than AcECT9 predominantly regulates m6A levels in ripening-related genes, thereby exerting the regulatory control over the ripening process and the accumulation of soluble sugars and organic acids, ultimately influencing fruit ripening and quality formation. In conclusion, our findings illuminate the epi-regulatory mechanism involving m6A in kiwifruit ripening, offering a fresh perspective for cultivating high-quality kiwifruit with enhanced nutritional attributes.

Equalization of RGB coupling efficiencies of metasurface waveguide coupler by adjusting imaginary part of refractive index.

Optical metasurfaces offer high-efficiency and flexible wavefront shaping for near-eye displays, especially in wideband waveguide couplers accommodating RGB primary colors. By leveraging the resonance characteristics of sub-wavelength periodic nanostructures, metasurfaces surpass the limitations of traditional optics that rely on multiple components and mediums. In this study, we propose adjustment of the imaginary parts of the material refractive indices as a new method to achieve balanced first-order diffraction efficiencies among RGB colors over a wide field of view (FOV) in an in-coupling metasurface waveguide coupler. Physical mechanism is investigated deeply and systematically in theory. It is found that nanostructure resonances deflect the wavefront and Poynting vector, significantly enhancing first-order diffraction efficiency, while resonance-enhanced absorption plays a crucial role in balancing the diffraction efficiency of RGB primary colors. First experimental demonstration well confirms the practical feasibility of this method and a uniform first-order diffraction efficiency of approximately 20% is achieved among RGB colors across a FOV as large as ∼30° over a single-piece glass substrate. This research provides insights into the design and mechanisms of metasurface waveguide couplers, advancing our understanding of metasurface-based RGB displays and facilitating further advancements in this field.

Multiplexed imaging in live cells using pulsed interleaved excitation spectral FLIM.

Multiplexed fluorescence detection has become increasingly important in the fields of biosensing and bioimaging. Although a variety of excitation/detection optical designs and fluorescence unmixing schemes have been proposed to allow for multiplexed imaging, rapid and reliable differentiation and quantification of multiple fluorescent species at each imaging pixel is still challenging. Here we present a pulsed interleaved excitation spectral fluorescence lifetime microscopic (PIE-sFLIM) system that can simultaneously image six fluorescent tags in live cells in a single hyperspectral snapshot. Using an alternating pulsed laser excitation scheme at two different wavelengths and a synchronized 16-channel time-resolved spectral detector, our PIE-sFLIM system can effectively excite multiple fluorophores and collect their emission over a broad spectrum for analysis. Combining our system with the advanced live-cell labeling techniques and the lifetime/spectral phasor analysis, our PIE-sFLIM approach can well unmix the fluorescence of six fluorophores acquired in a single measurement, thus improving the imaging speed in live-specimen investigation.