CRISPR-Cas9 Gene Editing for Sickle Cell Disease and ß-Thalassemia.

Transfusion-dependent ß-thalassemia (TDT) and sickle cell disease (SCD) are severe monogenic diseases with severe and potentially life-threatening manifestations. BCL11A is a transcription factor that represses γ-globin expression and fetal hemoglobin in erythroid cells. We performed electroporation of CD34+ hematopoietic stem and progenitor cells obtained from healthy donors, with CRISPR-Cas9 targeting the BCL11A erythroid-specific enhancer. Approximately 80% of the alleles at this locus were modified, with no evidence of off-target editing. After undergoing myeloablation, two patients - one with TDT and the other with SCD - received autologous CD34+ cells edited with CRISPR-Cas9 targeting the same BCL11A enhancer. More than a year later, both patients had high levels of allelic editing in bone marrow and blood, increases in fetal hemoglobin that were distributed pancellularly, transfusion independence, and (in the patient with SCD) elimination of vaso-occlusive episodes. (Funded by CRISPR Therapeutics and Vertex Pharmaceuticals; ClinicalTrials.gov numbers, NCT03655678 for CLIMB THAL-111 and NCT03745287 for CLIMB SCD-121.).

Controlling the confounding effect of metabolic gene expression to identify actual metabolite targets in microsatellite instability cancers.

BACKGROUND: The metabolome is the best representation of cancer phenotypes. Gene expression can be considered a confounding covariate affecting metabolite levels. Data integration across metabolomics and genomics to establish the biological relevance of cancer metabolism is challenging. This study aimed to eliminate the confounding effect of metabolic gene expression to reflect actual metabolite levels in microsatellite instability (MSI) cancers. METHODS: In this study, we propose a new strategy using covariate-adjusted tensor classification in high dimensions (CATCH) models to integrate metabolite and metabolic gene expression data to classify MSI and microsatellite stability (MSS) cancers. We used datasets from the Cancer Cell Line Encyclopedia (CCLE) phase II project and treated metabolomic data as tensor predictors and data on gene expression of metabolic enzymes as confounding covariates. RESULTS: The CATCH model performed well, with high accuracy (0.82), sensitivity (0.66), specificity (0.88), precision (0.65), and F1 score (0.65). Seven metabolite features adjusted for metabolic gene expression, namely, 3-phosphoglycerate, 6-phosphogluconate, cholesterol ester, lysophosphatidylethanolamine (LPE), phosphatidylcholine, reduced glutathione, and sarcosine, were found in MSI cancers. Only one metabolite, Hippurate, was present in MSS cancers. The gene expression of phosphofructokinase 1 (PFKP), which is involved in the glycolytic pathway, was related to 3-phosphoglycerate. ALDH4A1 and GPT2 were associated with sarcosine. LPE was associated with the expression of CHPT1, which is involved in lipid metabolism. The glycolysis, nucleotide, glutamate, and lipid metabolic pathways were enriched in MSI cancers. CONCLUSIONS: We propose an effective CATCH model for predicting MSI cancer status. By controlling the confounding effect of metabolic gene expression, we identified cancer metabolic biomarkers and therapeutic targets. In addition, we provided the possible biology and genetics of MSI cancer metabolism.

Functional characterization of stap2b in zebrafish vascular development.

The genetic control and signaling pathways of vascular development are not comprehensively understood. Transcription factors Islet2 (Isl2) and nr2f1b are critical for vascular growth in zebrafish, and further transcriptome analysis has revealed potential targets regulated by isl2/nr2f1b. In this study, we focused on the potential activation gene signal-transducing adaptor protein 2b (stap2b) and revealed a novel role of stap2b in vascular development. stap2b mRNA was expressed in developing vessels, suggesting stap2b plays a role in vascularization. Knocking down stap2b expression by morpholino injection or Crispr-Cas9-generated stap2b mutants caused vascular defects, suggesting a role played by stap2b in controlling the patterning of intersegmental vessels (ISVs) and the caudal vein plexus (CVP). The vessel abnormalities associated with stap2b deficiency were found to be due to dysregulated cell migration and proliferation. The decreased expression of vascular-specific markers in stap2b morphants was consistent with the vascular defects observed. In contrast, overexpression of stap2b enhanced the growth of ISVs and reversed the vessel defects in stap2b morphants. These data suggest that stap2b is necessary and sufficient to promote vascular development. Finally, we examined the interaction between stap2b and multiple signaling. We showed that stap2b regulated ISV growth through the JAK-STAT pathway. Moreover, we found that stap2b was regulated by Notch signaling to control ISV growth, and stap2b interacted with bone morphogenetic protein signaling to contribute to CVP formation. Altogether, we demonstrated that stap2b acts downstream of the isl2/nr2f1b pathway to play a pivotal role in vascular development via interaction with multiple signaling pathways.

Quantum pBac: An effective, high-capacity piggyBac-based gene integration vector system for unlocking gene therapy potential.

Recent advances in gene therapy have brought novel treatment options for cancer. However, the full potential of this approach has yet to be unlocked due to the limited payload capacity of commonly utilized viral vectors. Virus-free DNA transposons, including piggyBac, have the potential to obviate these shortcomings. In this study, we improved a previously modified piggyBac system with superior transposition efficiency. We demonstrated that the internal domain sequences (IDS) within the 3' terminal repeat domain of hyperactive piggyBac (hyPB) donor vector contain dominant enhancer elements. Plasmid-free donor vector devoid of IDS was used in conjunction with a helper plasmid expressing Quantum PBase™ v2 to generate an optimal piggyBac system, Quantum pBac™ (qPB), for use in T cells. qPB outperformed hyPB in CD20/CD19 CAR-T production in terms of performance as well as yield of the CAR-T cells produced. Furthermore, qPB also produced CAR-T cells with lower donor-associated variabilities compared to lentiviral vector. Importantly, qPB yielded mainly CD8+ CAR-TSCM cells, and the qPB-produced CAR-T cells effectively eliminated CD20/CD19-expressing tumor cells both in vitro and in vivo. Our findings confirm qPB as a promising virus-free vector system with an enhanced payload capacity to incorporate multiple genes. This highly efficient and potentially safe system will be expected to further advance gene therapy applications.

Engineered extracellular vesicles carrying let-7a-5p for alleviating inflammation in acute lung injury.

BACKGROUND: Acute lung injury (ALI) is a life-threatening respiratory condition characterized by severe inflammation and lung tissue damage, frequently causing rapid respiratory failure and long-term complications. The microRNA let-7a-5p is involved in the progression of lung injury, inflammation, and fibrosis by regulating immune cell activation and cytokine production. This study aims to use an innovative cellular electroporation platform to generate extracellular vesicles (EVs) carring let-7a-5p (EV-let-7a-5p) derived from transfected Wharton's jelly-mesenchymal stem cells (WJ-MSCs) as a potential gene therapy for ALI. METHODS: A cellular nanoporation (CNP) method was used to induce the production and release of EV-let-7a-5p from WJ-MSCs transfected with the relevant plasmid DNA. EV-let-7a-5p in the conditioned medium were isolated using a tangential flow filtration (TFF) system. EV characterization followed the minimal consensus guidelines outlined by the International Society for Extracellular Vesicles. We conducted a thorough set of therapeutic assessments, including the antifibrotic effects using a transforming growth factor beta (TGF-ß)-induced cell model, the modulation effects on macrophage polarization, and the influence of EV-let-7a-5p in a rat model of hyperoxia-induced ALI. RESULTS: The CNP platform significantly increased EV secretion from transfected WJ-MSCs, and the encapsulated let-7a-5p in engineered EVs was markedly higher than that in untreated WJ-MSCs. These EV-let-7a-5p did not influence cell proliferation and effectively mitigated the TGF-ß-induced fibrotic phenotype by downregulating SMAD2/3 phosphorylation in LL29 cells. Furthermore, EV-let-7a-5p regulated M2-like macrophage activation in an inflammatory microenvironment and significantly induced interleukin (IL)-10 secretion, demonstrating their modulatory effect on inflammation. Administering EVs from untreated WJ-MSCs slightly improved lung function and increased let-7a-5p expression in plasma in the hyperoxia-induced ALI rat model. In comparison, EV-let-7a-5p significantly reduced macrophage infiltration and collagen deposition while increasing IL-10 expression, causing a substantial improvement in lung function. CONCLUSION: This study reveals that the use of the CNP platform to stimulate and transfect WJ-MSCs could generate an abundance of let-7a-5p-enriched EVs, which underscores the therapeutic potential in countering inflammatory responses, fibrotic activation, and hyperoxia-induced lung injury. These results provide potential avenues for developing innovative therapeutic approaches for more effective interventions in ALI.

Clinical characteristics of membranous nephropathy after allogeneic hematopoietic stem cell transplantation: A real-world multicenter study.

Membranous nephropathy (MN) is a rare complication that can occur after allogeneic hematopoietic stem cell transplantation (allo-HSCT). MN patients may develop nephrotic syndrome or even kidney failure, which greatly affects their quality of life and prognosis. However, current knowledge regarding MN after allo-HSCT is limited. Thus, a multicenter nested case‒control study was conducted. Patients who had been diagnosed with MN after allo-HSCT were retrospectively identified at 8 HSCT centers. A total of 51 patients with MN after allo-HSCT were included. The median age of MN patients after allo-HSCT was 38 years, and the median duration from HSCT to MN was 18 months. The use of HLA-matched donors (P = 0.0102) and peripheral blood as the graft source (P = 0.0060) were identified as independent predisposing risk factors for the onset of MN after allo-HSCT. Compared to those in the control group, the incidence of extensive chronic graft-versus-host disease was greater in the MN patients (P = 0.0002). A total of 31 patients developed nephrotic syndrome. Patients receiving combination treatments of corticosteroids and immunosuppressants appeared to have better outcomes. In conclusion, MN is a rare but occasionally severe complication following HSCT and may require active treatment.

Niche and phenotypic differentiation in fern hybrid speciation, a case study of Pteris fauriei (Pteridaceae).

BACKGROUND AND AIMS: Niche differentiation is a crucial issue in speciation. Although it has a well-known role in adaptive processes of hybrid angiosperms, it is less understood in hybrid ferns. Here, we investigate whether an intermediate ecological niche of a fern hybrid is a novel adaptation that provides insights into fern hybrid speciation. METHODS: Pteris fauriei (Pteridaceae) is a natural hybrid fern, occurring in environments between its parent species. The maternal Pteris minor is found in sunny areas, but the habitat of the paternal Pteris latipinna is shady. We combined data from morphology, leaf anatomy and photosynthetic traits to explore adaptation and differentiation, along with measuring the environmental features of their niches. We also performed experiments in a common garden to understand ecological plasticity. KEY RESULTS: The hybrid P. fauriei was intermediate between the parent species in stomatal density, leaf anatomical features and photosynthetic characteristics in both natural habitats and a common garden. Interestingly, the maternal P. minor showed significant environmental plasticity and was more similar to the hybrid P. fauriei in the common garden, suggesting that the maternal species experiences stress in its natural habitats but thrives in environments similar to those of the hybrid. CONCLUSIONS: Based on the similar niche preferences of the hybrid and parents, we propose hybrid superiority. Our results indicate that the hybrid P. fauriei exhibits greater fitness and can compete with and occupy the initial niches of the maternal P. minor. Consequently, we suggest that the maternal P. minor has experienced a niche shift, elucidating the pattern of niche differentiation in this hybrid group. These findings offer a potential explanation for the frequent occurrence of hybridization in ferns and provide new insights into fern hybrid speciation, enhancing our understanding of fern diversity.

Uptake, translocation, and metabolism of organophosphate esters (OPEs) in plants and health perspective for human: A review.

Plant uptake, accumulation, and transformation of organophosphate esters (OPEs) play vital roles in their geochemical cycles and exposure risks. Here we reviewed the recent research advances in OPEs in plants. The mean OPE concentrations based on dry/wet/lipid weight varied in 4.80-3,620/0.287-26.8/12,000-315,000 ng g-1 in field plants, and generally showed positive correlations with those in plant habitats. OPEs with short-chain substituents and high hydrophilicity, particularly the commonly used chlorinated OPEs, showed dominance in most plant samples, whereas some tree barks, fruits, seeds, and roots demonstrated dominance of hydrophobic OPEs. Both hydrophilic and hydrophobic OPEs can enter plants via root and foliar uptake, and the former pathway is mainly passively mediated by various membrane proteins. After entry, different OPEs undergo diverse subcellular distributions and acropetal/basipetal/intergenerational translocations, depending on their physicochemical properties. Hydrophilic OPEs mainly exist in cell sap and show strong transferability, hydrophobic OPEs demonstrate dominant distributions in cell wall and limited migrations owing to the interception of Casparian strips and cell wall. Additionally, plant species, transpiration capacity, growth stages, commensal microorganisms, and habitats also affect OPE uptake and transfer in plants. OPE metabolites derived from various Phase I transformations and Phase II conjugations are increasingly identified in plants, and hydrolysis and hydroxylation are the most common metabolic processes. The metabolisms and products of OPEs are closely associated with their structures and degradation resistance and plant species. In contrast, plant-derived food consumption contributes considerably to the total dietary intakes of OPEs by human, particularly the cereals, and merits specifical attention. Based on the current research limitations, we proposed the research perspectives regarding OPEs in plants, with the emphases on their behavior and fate in field plants, interactions with plant-related microorganisms, multiple uptake pathways and mechanisms, and comprehensive screening analysis and risk evaluation.

Effects of down-regulated carbonic anhydrase 8 on cell survival and glucose metabolism in human colorectal cancer cell lines.

Carbonic anhydrase 8 (CA8) is a member of the α-carbonic anhydrase family but does not catalyze the reversible hydration of carbon dioxide. In the present study, we examined the effects of CA8 on two human colon cancer cell lines, SW480 and SW620, by suppressing CA8 expression through shRNA knockdown. Our results showed that knockdown of CA8 decreased cell growth and cell mobility in SW620 cells, but not in SW480 cells. In addition, downregulated CA8 resulted in a significant decrease of glucose uptake in both SW480 and SW620 cells. Interestingly, stable downregulation of CA8 decreased phosphofructokinase-1 expression but increased glucose transporter 3 (GLUT3) levels in SW620 cells. However, transient downregulation of CA8 fails to up-regulate GLUT3 expression, indicating that the increased GLUT3 observed in SW620-shCA8 cells is a compensatory effect. In addition, the interaction between CA8 and GLUT3 was evidenced by pull-down and IP assays. On the other hand, we showed that metformin, a first-line drug for type II diabetes patients, significantly inhibited cell migration of SW620 cells, depending on the expressions of CA8 and focal adhesion kinase. Taken together, our data demonstrate that when compared to primary colon cancer SW480 cells, metastatic colon cancer SW620 cells respond differently to downregulated CA8, indicating that CA8 in more aggressive cancer cells may play a more important role in controlling cell survival and metformin response. CA8 may affect glucose metabolism- and cell invasion-related molecules in colon cancer, suggesting that CA8 may be a potential target in future cancer therapy.

Turn-off enzyme activity of histidine-rich peptides for the detection of lysozyme.

An assay that integrates histidine-rich peptides (HisRPs) with high-affinity aptamers was developed enabling the specific and sensitive determination of the target lysozyme. The enzyme-like activity of HisRP is inhibited by its interaction with a target recognized by an aptamer. In the presence of the target, lysozyme molecules progressively assemble on the surface of HisRP in a concentration-dependent manner, resulting in the gradual suppression of enzyme-like activity. This inhibition of HisRP's enzyme-like activity can be visually observed through color changes in the reaction product or quantified using UV-visible absorption spectroscopy. Under optimal conditions, the proposed colorimetric assay for lysozyme had a detection limit as low as 1 nM and exhibited excellent selectivity against other nonspecific interferents. Furthermore, subsequent research validated the practical applicability of the developed colorimetric approach to saliva samples, indicating that the assay holds significant potential for the detection of lysozymes in samples derived from humans.

[Cloning and functional analysis of flavanone synthase â ¡ from Dendrobium huoshanense].

Flavonoid C-glycosides are a class of natural products that are widely involved in plant defense responses and have diverse pharmacological activities. They are also important active ingredients of Dendrobium huoshanense. Flavanone synthase Ⅱ has been proven to be a key enzyme in the synthesis pathway of flavonoid C-glycosides in plants, and their catalytic product 2-hydroxyflavanone is the precursor compound for the synthesis of various reported flavonoid C-glycosides. In this study, based on the reported amino acid sequence of flavanone synthase Ⅱ, a flavanone synthase Ⅱ gene(DhuFNSⅡ) was screened and verified from the constructed D. huoshanense genome localization database. Functional validation of the enzyme showed that it could in vitro catalyze naringenin and pinocembrin to produce apigenin and chrysin, respectively. The open reading frame(ORF) of DhuFNSⅡ was 1 644 bp in length, encoding 547 amino acids. Subcellular localization showed that the protein was localized on the endoplasmic reticulum. RT-qPCR results showed that DhuFNSⅡ had the highest expression in stems, followed by leaves and roots. The expression levels of DhuFNSⅡ and other target genes in various tissues of D. huoshanense were significantly up-regulated after four kinds of abiotic stresses commonly encountered in the growth process, but the extent of up-regulation varied among treatment groups, with drought and cold stress having more significant effects on gene expression levels. Through the identification and functional analysis of DhuFNSⅡ, this study is expected to contribute to the elucidation of the molecular mechanism of the formation of quality metabolites of D. huoshanense, flavonoid C-glycosides, and provide a reference for its quality formation and scientific cultivation.

Sandwich Structured Metal oxide/Reduced Graphene Oxide/Metal Oxide-Based Polymer Electrolyte Enables Continuous Inorganic-Organic Interphase for Fast Lithium-Ion Transportation.

Introducing inorganic fillers into organic poly(ethylene oxide)(PEO)-based electrolyte has attracted substantial attention to enhance its ionic conductivity and mechanical strength, but limited inorganic-organic interphases are always caused by isolated particles agglomeration. Herein, a variety of sandwich structured metal oxide/reduced graphene oxide(rGO)/metal oxide nanocomposites to optimize lithium-ion conduction by interconnected amorphous organic-inorganic interphases in lithium metal batteries, are proposed. With the support of high surface area rGO, the agglomeration of metal oxide particles is precluded, forming continuous amorphous organic-inorganic interphases with stacked layer-by-layer structure, thus creating 3D interconnected lithium-ion transportation channels vertically and laterally. Besides, metal oxide nanoparticles with hydroxyls possess high affinity toward bis(tri-fluoromethanesulfonyl)imide anions by hydrogen bindings between hydroxyls and fluorine and metal-oxygen bonds, releasing more free lithium ions. Consequently, PEO-ZnO/rGO/ZnO electrolyte delivers superior ionic conductivity of 1.02 × 10-4 S cm-1 at 25 °C and lithium-ion transference number of 0.38 at 60 °C. Furthermore, ZnO/rGO/ZnO insertion promotes the formation of LiF-rich stable solid electrolyte interface, endowing Li symmetric cells with long-term cycling stability over 900 hours. The corresponding LiFePO4 cathode possesses a high reversible specific capacity of 130 mAh g-1 at 0.5C after cycling 300 cycles with a poor capacity fading of 0.05% per cycle.

Immunodeficiency and bone marrow failure with mosaic and germline TLR8 gain of function.

Inborn errors of immunity (IEI) are a genetically heterogeneous group of disorders with a broad clinical spectrum. Identification of molecular and functional bases of these disorders is important for diagnosis, treatment, and an understanding of the human immune response. We identified 6 unrelated males with neutropenia, infections, lymphoproliferation, humoral immune defects, and in some cases bone marrow failure associated with 3 different variants in the X-linked gene TLR8, encoding the endosomal Toll-like receptor 8 (TLR8). Interestingly, 5 patients had somatic variants in TLR8 with <30% mosaicism, suggesting a dominant mechanism responsible for the clinical phenotype. Mosaicism was also detected in skin-derived fibroblasts in 3 patients, demonstrating that mutations were not limited to the hematopoietic compartment. All patients had refractory chronic neutropenia, and 3 patients underwent allogeneic hematopoietic cell transplantation. All variants conferred gain of function to TLR8 protein, and immune phenotyping demonstrated a proinflammatory phenotype with activated T cells and elevated serum cytokines associated with impaired B-cell maturation. Differentiation of myeloid cells from patient-derived induced pluripotent stem cells demonstrated increased responsiveness to TLR8. Together, these findings demonstrate that gain-of-function variants in TLR8 lead to a novel childhood-onset IEI with lymphoproliferation, neutropenia, infectious susceptibility, B- and T-cell defects, and in some cases, bone marrow failure. Somatic mosaicism is a prominent molecular mechanism of this new disease.

Benefit of [18F] FDG PET/CT in the diagnosis and salvage treatment of recurrent nasopharyngeal carcinoma.

PURPOSE: To compare PET/CT, MRI and ultrasonography in detecting recurrence of nasopharyngeal carcinoma and identify their benefit in staging, contouring and overall survival (OS). METHODS: Cohort A included 1453 patients with or without histopathology-confirmed local recurrence, while cohort B consisted of 316 patients with 606 histopathology-confirmed lymph nodes to compare the sensitivities and specificities of PET/CT, MRI and ultrasonography using McNemar test. Cohorts C and D consisted of 273 patients from cohort A and 267 patients from cohort B, respectively, to compare the distribution of PET/CT-based and MRI-based rT-stage and rN-stage and the accuracy of rN-stage using McNemar test. Cohort E included 30 random patients from cohort A to evaluate the changes in contouring with or without PET/CT by related-samples T test or Wilcoxon rank test. The OS of 61 rT3-4N0M0 patients staged by PET/CT plus MRI (cohort F) and 67 MRI-staged rT3-4N0M0 patients (cohort G) who underwent similar salvage treatment were compared by log-rank test and Cox regression. RESULTS: PET/CT had similar specificity to MRI but higher sensitivity (93.9% vs. 79.3%, P < 0.001) in detecting local recurrence. PET/CT, MRI and ultrasonography had comparable specificities, but PET/CT had greater sensitivity than MRI (90.9% vs. 67.6%, P < 0.001) and similar sensitivity to ultrasonography in diagnosing lymph nodes. According to PET/CT, more patients were staged rT3-4 (82.8% vs. 68.1%, P < 0.001) or rN + (89.9% vs. 69.3%, P < 0.001), and the rN-stage was more accurate (90.6% vs. 73.8%, P < 0.001). Accordingly, the contours of local recurrence were more precise (median Dice similarity coefficient 0.41 vs. 0.62, P < 0.001) when aided by PET/CT plus MRI. Patients staged by PET/CT plus MRI had a higher 3-year OS than patients staged by MRI alone (85.5% vs. 60.4%, P = 0.006; adjusted HR = 0.34, P = 0.005). CONCLUSION: PET/CT more accurately detected and staged recurrence of nasopharyngeal carcinoma and accordingly complemented MRI, providing benefit in contouring and OS.

A non-genetic engineering platform for rapidly generating and expanding cancer-specific armed T cells.

BACKGROUND: Cancer-specific adoptive T cell therapy has achieved successful milestones in multiple clinical treatments. However, the commercial production of cancer-specific T cells is often hampered by laborious cell culture procedures, the concern of retrovirus-based gene transfection, or insufficient T cell purity. METHODS: In this study, we developed a non-genetic engineering technology for rapidly manufacturing a large amount of cancer-specific T cells by utilizing a unique anti-cancer/anti-CD3 bispecific antibody (BsAb) to directly culture human peripheral blood mononuclear cells (PBMCs). The anti-CD3 moiety of the BsAb bound to the T cell surface and stimulated the differentiation and proliferation of T cells in PBMCs. The anti-cancer moiety of the BsAb provided these BsAb-armed T cells with the cancer-targeting ability, which transformed the naïve T cells into cancer-specific BsAb-armed T cells. RESULTS: With this technology, a large amount of cancer-specific BsAb-armed T cells can be rapidly generated with a purity of over 90% in 7 days. These BsAb-armed T cells efficiently accumulated at the tumor site both in vitro and in vivo. Cytotoxins (perforin and granzyme) and cytokines (TNF-α and IFN-γ) were dramatically released from the BsAb-armed T cells after engaging cancer cells, resulting in a remarkable anti-cancer efficacy. Notably, the BsAb-armed T cells did not cause obvious cytokine release syndrome or tissue toxicity in SCID mice bearing human tumors. CONCLUSIONS: Collectively, the BsAb-armed T cell technology represents a simple, time-saving, and highly safe method to generate highly pure cancer-specific effector T cells, thereby providing an affordable T cell immunotherapy to patients.

Identification and characterization of two long-type peptidoglycan recognition proteins, PGRP-L1 and PGRP-L2, in the orange-spotted grouper, Epinephelus coioides.

Peptidoglycan recognition proteins (PGRPs) play an important role in innate immunity by recognizing components of pathogenic bacteria (such as peptidoglycan, PGN) and are evolutionarily conserved pattern recognition receptors (PRRs) in both invertebrates and vertebrates. In the present study, two long-type PGRPs (designed as Eco-PGRP-L1 and Eco-PGRP-L2) were identified in orange-spotted grouper (Epinephelus coioides), which is a major economic species cultured in Asia. The predicted protein sequences of both Eco-PGRP-L1 and Eco-PGRP-L2 contain a typical PGRP domain. Eco-PGRP-L1 and Eco-PGRP-L2 exhibited organ/tissue-specific expression patterns. An abundant expression of Eco-PGRP-L1 was observed in pyloric caecum, stomach and gill, whereas a highest expression level of Eco-PGRP-L2 was found in head kidney, spleen, skin and heart. In addition, Eco-PGRP-L1 is distributed in the cytoplasm and nucleus, while Eco-PGRP-L2 is mainly localized in cytoplasm. Both Eco-PGRP-L1 and Eco-PGRP-L2 were induced following the stimulation of PGN and have PGN binding activity. In addition, functional analysis revealed that Eco-PGRP-L1 and Eco-PGRP-L2 possess antibacterial activity against Edwardsiella tarda. These results may contribute to understand the innate immune system of orange-spotted grouper.

Predicting in-hospital mortality risk for perforated peptic ulcer surgery: the PPUMS scoring system and the benefit of laparoscopic surgery: a population-based study.

BACKGROUND: The major treatment for perforated peptic ulcers (PPU) is surgery. It remains unclear which patient may not get benefit from surgery due to comorbidity. This study aimed to generate a scoring system by predicting mortality for patients with PPU who received non-operative management (NOM) and surgical treatment. METHOD: We extracted the admission data of adult (≥ 18 years) patients with PPU disease from the NHIRD database. We randomly divided patients into 80% model derivation and 20% validation cohorts. Multivariate analysis with a logistic regression model was applied to generate the scoring system, PPUMS. We then apply the scoring system to the validation group. RESULT: The PPUMS score ranged from 0 to 8 points, composite with age (< 45: 0 points, 45-65: 1 point, 65-80: 2 points, > 80: 3 points), and five comorbidities (congestive heart failure, severe liver disease, renal disease, history of malignancy, and obesity: 1 point each). The areas under ROC curve were 0.785 and 0.787 in the derivation and validation groups. The in-hospital mortality rates in the derivation group were 0.6% (0 points), 3.4% (1 point), 9.0% (2 points), 19.0% (3 points), 30.2% (4 points), and 45.9% when PPUMS > 4 point. Patients with PPUMS > 4 had a similar in-hospital mortality risk between the surgery group [laparotomy: odds ratio (OR) = 0.729, p = 0.320, laparoscopy: OR = 0.772, p = 0.697] and the non-surgery group. We identified similar results in the validation group. CONCLUSION: PPUMS scoring system effectively predicts in-hospital mortality for perforated peptic ulcer patients. It factors in age and specific comorbidities is highly predictive and well-calibrated with a reliable AUC of 0.785-0.787. Surgery, no matter laparotomy or laparoscope, significantly reduced mortality for scores < = 4. However, patients with a score > 4 did not show this difference, calling for tailored approaches to treatment based on risk assessment. Further prospective validation is suggested.

Impact of age-friendly living environment and intrinsic capacity on functional ability in older adults: a cross-sectional study.

BACKGROUND: The World Health Organization (WHO) has proposed healthy aging framework, supposing that intrinsic capacity (IC), environment and their interaction may have influence on functional ability (FA). It was still unclear how the IC level and age-friendly living environment impact on FA. This study aims to confirm the relationship between the IC level and age-friendly living environment with FA, especially in older adults with low IC. METHODS: Four hundred eighty-five community-dwelling residents aged ≥ 60 years were enrolled. IC constructed by locomotion, cognition, psychological, vitality, and sensory domains was assessed using full assessment tools recommended by WHO. Age-friendly living environment was measured with 12 questions adapted from the spatial indicators framework of age-friendly cities. FA was assessed using activities of daily living (ADL) and one question about mobile payment ability. Multivariate logistic regression was used to explore the association between IC, environment and FA. The influence of the environment on electronic payment and ADL under the IC layer was assessed. RESULTS: Of 485 respondents, 89 (18.4%) had ADL impairment, and 166 (34.2%) had mobile payment function impairment. Limited IC (odds ratio [OR] = 0.783, 95% confidence interval [CI] = 0.621-0.988) and poor environment (OR = 0.839, 95% CI = 0.733-0.960) were associated with mobile payment ability impairment. Our results suggested that a supportive age-friendly living environment influenced FA was more prominent in older adults with poor IC (OR = 0.650, 95% CI = 0.491-0.861). CONCLUSIONS: Our results confirmed IC and the environment had an impact on mobile payment ability. The relationship between environment and FA showed differences according to IC level. These findings suggest that an age-friendly living environment is important to maintain and enhance elders' FA, especially in those with poor IC.

A strategy combining chemical analysis and network pharmacology to investigate the mechanism of Xiao'er Qingre Zhike Oral solution in cough.

Xiao'er Qingre Zhike Oral Solution (XQZS) is a commonly used TCM formula to treat cough in children in China. Its complicated composition renders its chemical analysis and mechanism elucidation difficult. To evaluate the bioactive components and mechanism of XQZS against cough, we used a combination strategy of chemical analysis and network pharmacology. A UHPLC/Q-Orbitrap-MS method was established for the identification and qualitative analysis of components of XQZS, and a total of 33 components were unambiguously identified. Aiming at identifying the components, network pharmacology revealed 107 potential targets related to cough. Using protein-protein interactions analysis, nine core targets were selected. Several cough-related pathways were enriched using the Kyoto Encyclopedia of Genes and Genomes, including neuroactive ligand-receptor interaction, serotonergic synapse and dopaminergic synapse. The herb-compound-target-pathway network indicated that PTGS2 (COX-2) was the core target of XQZS against cough. To demonstrate the inhibition effects of the major components against the key target, a COX-2 inhibitor screening assay was used. Compounds P2, P4, P23 and P49 exhibited promising inhibition effects on COX-2 at 20 µm, with inhibitory rates of 55.80-69.87%. In conclusion, this study demonstrates that XQZS may alleviate cough via the inhibition of PTGS2 (COX-2) and the regulation of the serotonergic synapse pathway. The chemical analysis and network pharmacology integrated evaluation provided an efficient strategy for discovering the key pharmacological mechanism of XQZS.

Association between cumulative duration of deep anesthesia and postoperative acute kidney injury after noncardiac surgeries: a retrospective observational study.

BACKGROUND: Bispectral index (BIS) is a processed electroencephalography monitoring tool and is widely used in anesthetic depth monitoring. Deep anesthesia exposure may be associated with multiple adverse outcomes. However, the relationship between anesthetic depth and postoperative acute kidney injury (AKI) remains unclear. We sought to determine the effect of BIS-based deep anesthesia duration on postoperative AKI following noncardiac surgery. METHODS: This retrospective study used data from the Vital Signs DataBase, including patients undergoing noncardiac surgeries with BIS monitoring. The BIS values were collected every second during anesthesia. Restricted cubic splines and logistic regression were used to assess the association between the cumulative duration of deep anesthesia and postoperative AKI. RESULTS: 4774 patients were eligible, and 129 (2.7%) experienced postoperative AKI. Restricted cubic splines showed that a cumulative duration of BIS < 45 was nonlinearly associated with postoperative AKI (P-overall = 0.033 and P-non-linear = 0.023). Using the group with the duration of BIS < 45 less than 15 min as the reference, ORs of postoperative AKI were 2.59 (95% confidence interval [CI]:0.60 to 11.09, p = 0.200) in the 15-100 min group, and 4.04 (95%CI:0.92 to 17.76, p = 0.064) in the ≥ 100 min group after adjusting for preoperative and intraoperative covariates in multivariable logistic regression. CONCLUSIONS: The cumulative duration of BIS < 45 was independently and nonlinearly associated with the risk of postoperative AKI in patients undergoing noncardiac surgery.