Validation of hyperthermia as an enhancer of chemotherapeutic efficacy: insights from a bladder cancer organoid model.

OBJECTIVE: This study aimed to evaluate the combined efficacy of hyperthermia and chemotherapy using a bladder cancer organoid model and to explore hyperthermia-related molecular pathways. METHOD: Tumor organoids were generated by embedding RT4 bladder cancer cells into Matrigel. The resulting organoids were treated with pirarubicin or gemcitabine at 37 °C or 42 °C. Proliferation was determined by Ki67 immunofluorescence staining, and apoptosis was assessed using a TdT-mediated dUTP nick end labeling (TUNEL) assay. RNA sequencing was used to identify the differentially expressed genes. RESULTS: Bladder cancer organoids were successfully established and exhibited robust proliferative abilities. Treatment with gemcitabine or pirarubicin under hyperthermic conditions caused pronounced structural damage to the organoids and increased cell death compared to that in the normothermically treated group. Furthermore, Ki67 labeling and TUNEL assays showed that the hyperthermia chemotherapy group showed a significantly reduced proliferation rate and high level of apoptosis. Finally, RNA sequencing revealed the IFN-γ signaling pathway to be associated with hyperthermia. CONCLUSION: Overall, hyperthermia combined with chemotherapy exerted better therapeutic effects than those of normothermic chemotherapy in grade 1-2 non-muscle-invasive bladder cancer, potentially through activation of the IFN-γ-JAK-STAT pathway.

Effect of Esketamine Gargle on Postoperative Sore Throat in Patients Undergoing Double-Lumen Endobronchial Intubation: A Randomised Controlled Trial.

Background: Postoperative sore throat is a frequent adverse event after double-lumen endobronchial tube (DLT) intubation. The aim of this study was to investigate whether esketamine gargle has a preventive effect on the incidence of postoperative sore throat in patients undergoing DLT intubation. Methods: This trial included 140 patients undergoing elective thoracic surgery at the third affiliated hospital of Sun Yat-Sen University. Patients were randomly allocated into the following two groups of 70 patients each: the control group, gargling with saline of 30 mL, and the esketamine group, gargling with an esketamine solution of 30 mL (2 mL/50 mg esketamine in 28 mL saline), 5 min prior to anaesthesia induction. The primary outcome was the incidence of sore throat 24 h after surgery. The main secondary outcomes included the incidence of sore throat and hoarseness at 1 h and 48 h after surgery, as well as the intraoperative haemodynamic responses. Results: The incidence of sore throat was significantly higher in the control group (47.1%, 33/70 patients) compared with the esketamine group (12.9%, 9/70 patients) at 24 h after surgery (RD, 0.41; 95% confidence interval, 0.26-0.57; p < 0.001). Furthermore, the incidence of sore throat at 1 h (p = 0.027), 24 h (p = 0.019), and seventh day (p = 0.031) as well as hoarseness at 1 h (p = 0.027), 24 h (p = 0.019), and 48 h (p = 0.031) after surgery were reduced in the esketamine group. Significant differences were seen in the peak levels of systolic blood pressure, mean arterial blood pressure, and heart rate between the groups during the intubation (p < 0.05). Besides, no patient developed an adverse reaction to esketamine. Conclusion: Preoperative gargling of esketamine can provide an effect against sore throat after DLT intubation without adverse side effects and effectively inhibit sharp elevations in heart rate and blood pressure during double-lumen intubation procedures.

Molecular docking and dynamics of a dextranase derived from Penicillium cyclopium CICC-4022.

This study aimed to investigate the recognition mechanism of dextranase (PC-Edex) produced by Penicillium cyclopium CICC-4022 on dextran. Whole genome information of P. cyclopium CICC-4022 was obtained through genome sequencing technology. The coding information of PC-Edex was determined based on the annotation of the protein-coding genes using protein databases. The three-dimensional structure of PC-Edex was obtained via homology modelling. The active site and binding free energy between PC-Edex and dextran were calculated by molecular docking and molecular dynamics techniques. The results showed that the total sequence length and GC content of P. cyclopium CICC-4022 were 29,710,801 bp and 47.02 %, respectively. The annotation of protein-encoding genes showed that P. cyclopium CICC-4022 is highly active and has many carbohydrate transport and metabolic functions, and most of its proteases are glycolytic anhydrases. Furthermore, the gene encoding PC-Edex was successfully annotated. Molecular dynamics simulations indicated that van der Waals interaction was the main driving force of interaction. Residues Ile114, Asp115, Tyr332, Lys344, and Gln403 significantly promoted the binding between dextran and PC-Edex. In summary, this study explored the active site catalyzed by PC-Edex based on the binding pattern of PC-Edex and dextran. Therefore, this study provides genomic information on dextranase and data supporting the rational modification and enhancement of PC-Edex.

Loss of SETD2-mediated downregulation of intracellular and exosomal miRNA-10b determines MAPK pathway activation and multidrug resistance in renal cancer.

SET domain-containing 2 (SETD2) is the most frequently mutated gene among all the histone methyltransferases in clear cell renal cell carcinoma (ccRCC). Microarrays, RNA sequencing analysis and exosomes analysis of cellular supernatant were performed after transfection A498 cells with si-SETD2 or siRNA of negative control. Chromatin immunoprecipitation and Luciferase reporter assay were conducted to evaluate the interaction between SETD2 and miR-10b. Functional and drug experiments in vitro and in vivo were performed to verify the role of SETD2, miR-10b and MAP4K4. The results showed that loss of SETD2 mediated downregulation of intracellular and exosomal microRNA-10b. MAP4K4 were relevant to oncogenesis of ccRCC caused by loss of SETD2 and miR-10b. SETD2 could directly target miR-10b and regulate the expression of multidrug resistance (MDR)-1 (P-gp170) through JNK pathway, which was one of the downstream pathways of MAP4K4. The coordinated expression of SETD2/H3K36me3/miR-10b/MAPKs/JNK/MDR pathway was revealed to the progression of ccRCC.

Downregulation of connexin 43-based gap junctions underlies propofol-induced excessive relaxation in hypertensive vascular smooth muscle cells.

BACKGROUND: Postinduction hypotension caused by propofol remains a non-negligible problem for anesthesiologists, and is especially severe in chronic hypertensive patients with long-term vasoconstriction and decreased vascular elasticity. The functional change in gap junctions composed of Cx43 (Cx43-GJs) is reported as the biological basis of synchronized contraction or relaxation of blood vessels. Thus, we investigated the role of Cx43-GJs in propofol-induced dramatic blood pressure fluctuations in chronic hypertensive patients, and their internal mechanisms. METHODS: Human umbilical artery smooth muscle cells (HUASMCs) were pretreated with long-term angiotensin II (Ang II), with or without propofol, to simulate the contraction and relaxation of normal and hypertensive VSMCs during anesthesia induction. The levels of F-actin polymerization and MLC2 phosphorylation were used as indicators to observe the contraction and relaxation of HUASMCs. Different specific activators, inhibitors and siRNAs were used to explore the role of Cx43-GJs and Ca2+ as well as the RhoA/ LIMK2/cofilin and RhoA/MLCK signaling pathways in the contraction and relaxation of normal and hypertensive HUASMCs. RESULTS: Both F-actin polymerization and MLC2 phosphorylation were significantly enhanced in Ang II-pretreated HUASMCs, along with higher expression of Cx43 protein and stronger function of Cx43-GJs than in normal HUASMCs. However, with propofol administration, similar to Gap26 and Cx43-siRNA, the function of Cx43-GJs in Ang II-pretreated HUASMCs was inhibited compared with that in normal HUASMCs, accompanied by a larger decrease in intracellular Ca2+ and the RhoA/LIMK2/cofilin and RhoA/MLCK signaling pathways. Eventually F-actin polymerization and MLC2 phosphorylation were more dramatically decreased. However, these effects could be reversed by RA with enhanced Cx43-GJ function. CONCLUSION: Long-term exposure to Ang II significantly enhanced the expression of the Cx43 protein and function of Cx43-GJs in HUASMCs, resulting in the accumulation of intracellular Ca2+ and the activation of its downstream RhoA/LIMK2/cofilin and RhoA/MLCK signaling pathways, which maintained HUASMCs in a state of excessive-contraction. With inhibition of Cx43-GJs by propofol in Ang II-pretreated HUASMCs, intracellular Ca2+ and its downstream signaling pathways were dramatically inhibited, which ultimately excessively relaxed HUASMCs. This is the reason why the blood pressure fluctuation of patients with chronic hypertension was more severe after receiving propofol induction. Video Abstract.

Effects of Dietary Protein and Lipid Levels on Growth, Metabolism, Antioxidative Capacity, and Fillet Quality of Adult Triploid Rainbow Trout Farmed in Net Cage.

The research is aimed at investigating the effects of dietary protein and lipid levels on adult triploid rainbow trout growth performance, feed utilization, digestive and metabolic enzyme activities, antioxidative capacity, and fillet quality. Nine diets containing three dietary protein levels (DP) (300, 350, and 400 g kg-1) and three dietary lipid levels (DL) (200, 250, and 300 g kg-1) were prepared using a 3 × 3 factorial design. In freshwater cages, 13,500 adult female triploid rainbow trout (3.2 ± 0.1 kg) were cultured for 77 days. Triplicate cages (500 fish per cage) were used as repetitions of each experimental diet. The findings revealed that as DP increased to 400 g kg-1 and DL raised to 300 g kg-1, the weight gain ratio (WGR) elevated significantly (P < 0.05). However, when DP ≥ 350 g kg-1, WGR was similar in the DL250 and DL300 groups. As DP raised to 350 g kg-1, the feed conversion ratio (FCR) notably decreased (P < 0.05). In the DP350DL300 group, lipids had a protein-sparing impact. High DP diet (400 g kg-1) generally improved fish health status by increasing antioxidant capacity in the liver and intestine. A high DL diet (300 g kg-1) showed no harmful effect on hepatic health based on plasma levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) and antioxidant capacity in the liver. For fillet quality, a high DP diet could increase fillet yield, improve fillet hardness, springiness, and water-holding capacity values, and inhibit the production of off-flavors caused by n-6 fatty acids. A high DL diet could increase odor intensity, and EPA, DHA, and n-3 fatty acid concentrations decrease the thrombogenicity index value. The maximum fillet redness value was discovered in the DP400DL300 group. Overall, for adult triploid rainbow trout (≥3 kg), the minimum recommended DP and DL according to growth performance were 400 and 250 g kg-1, respectively; DP and DL based on feed utilization were 350 and 200 g kg-1, respectively; DP and DL based on fillet quality were 400 and 300 g kg-1, respectively.

A spatiotemporally controlled recombinant cccDNA mouse model for studying HBV and developing drugs against the virus.

Covalently closed circular (ccc) DNA is the template for hepatitis B virus (HBV) replication. The lack of small animal models for characterizing chronic HBV infection has hampered research progress in HBV pathogenesis and drug development. Here, we generated a spatiotemporally controlled recombinant cccDNA (rcccDNA) mouse model by combining Cre/loxP-mediated DNA recombination with the liver-specific "Tet-on/Cre" system. The mouse model harbors three transgenes: a single copy of the HBV genome (integrated at the Rosa26 locus, RHBV), H11-albumin-rtTA (spatiotemporal conditional module), and (tetO)7-Cre (tetracycline response element), and is named as RHTC mouse. By supplying the RHTC mice with doxycycline (DOX)-containing drinking water for two days, the animals generate rcccDNA in hepatocytes, and the rcccDNA supports active HBV gene expression and can maintain HBV viremia persistence for over 60 weeks. Persistent HBV gene expression induces intrahepatic inflammation, fibrosis, and dysplastic pathology, which closely mirrors the disease progression in clinical patients. Bepirovirsen, an antisense oligonucleotide (ASO) targeting all HBV RNA species, showed dose-dependent antiviral effects in the RHTC mouse model. The spatiotemporally controlled rcccDNA mouse is convenient and reliable, providing versatile small animal model for studying cccDNA-centric HBV biology as well as evaluating antiviral therapeutics.

Adult Triploid Rainbow Trout Can Adapt to Various Dietary Lipid Levels by Coordinating Metabolism in Different Tissues.

Triploid rainbow trout can adapt to various dietary lipid levels; however, the mechanisms of systematic adaptation are not well understood. To investigate how adult triploid rainbow trout maintains lipid hemostasis under different exogenous lipid intake, a 77-day feeding trial was conducted. Diets with lipid contents of 20%, 25%, and 30% were formulated and fed to triploid rainbow trout with an initial weight of 3 ± 0.02 kg, and they were named L20, L25, and L30 group, respectively. Results showed that the condition factor, hepatosomatic index, liver color, and plasma triglyceride were comparable among three groups (p > 0.05), whereas the value of specific growth rate, viscerosomatic index, and liver glycogen content gradually increased with increasing dietary lipid level (p < 0.05). A significantly highest value of plasma glucose and nonesterified fatty acids were found in the L30 group (p < 0.05), whereas the significantly higher content of plasma total cholesterol, high-density lipoprotein-cholesterol, and low-density lipoprotein-cholesterol was found in the L25 group compared with those in L20 group (p < 0.05). As for lipid deposition, abdominal adipose tissue, and muscle were the main lipid storage place for triploid rainbow trout when tissues' weight is taken into consideration. Overall quantitative PCR showed that the lipid transport and glycolysis were upregulated, and fatty acids oxidative was downregulated in liver when fish were fed low lipid diets. It meant that the liver was the primary lipid metabolizing organ to low lipid diet feeding, which could switch energy supply between glycolysis and fatty acids oxidation. Fish fed with a moderate dietary lipid level diet could increase lipid uptake and promote lipogenesis in muscle. Abdominal adipose tissue could efficiently uptake excess exogenous free fatty acid through upregulating fatty acid uptake and synthesis de novo and then storing it in the form of triglyceride. Excess lipid uptake is preferentially stored in abdominal adipose tissue through coordinated fatty acid uptake and fatty acid synthesis de novo as dietary lipid levels increased. In summary, triploid rainbow trout can adapt to various dietary lipid levels by coordinating metabolism in different tissues.

Corrigendum: Transcription factors BARX1 and DLX4 contribute to progression of clear cell renal cell carcinoma via promoting proliferation and epithelial-mesenchymal transition.

[This corrects the article DOI: 10.3389/fmolb.2021.626328.].

[Research and Application of Seismic Performance of Medical Imaging Equipment].

In view of the shortage of research on the seismic performance of medical imaging equipment, this paper investigates and summarizes the seismic regulatory requirements and seismic tests of medical imaging equipment, and focuses on the parameter selection, detection steps, result evaluation and detection equipment requirements of seismic detection of medical imaging equipment. The seismic test data of medical imaging equipment with various installation modes are analyzed, and the seismic performance of medical imaging equipment is analyzed and summarized.

A Case Report of Adrenal Teratoma: Huge but Easy to Misdiagnose.

Primary adrenal teratoma is a rare lesion with a high misdiagnosis rate. In view of the fact that adrenal teratoma and adrenal gland are often inseparable, the removal of the affected adrenal gland seems inevitable. Thus, it is necessary for clinicians to differentiate adrenal teratoma from common adrenal lipoma diseases, such as adrenal angiomyolipoma, adrenal myelolipoma, lipoma, gangliocytoma, and so on. In this study, we reported the clinical and pathologic features of 7 patients surgically treated for adrenal mature teratomas from 2007 to 2020 in our hospital. We compared the clinical characteristics and surgical management in literature review cases with our cases. All patients were females and underwent open adrenalectomy or laparoscopic adrenalectomy, respectively. In our cases, there was no recurrence during the longest follow-up of 13 years. Adrenal teratoma is rare and easy to ignore. And once misdiagnosed, the surgeon may need to adjust the operation temporarily, extend the operation time and finally increase the operative risks. Our aim is mainly to help clinicians to raise awareness of adrenal teratomas, improve the diagnosis rates and optimize the treatment decision-making of this disease.

Injectable electrospun fiber-hydrogel composite sequentially releasing clonidine and ropivacaine for prolonged and walking regional analgesia.

Rationale: Peripheral nerve block is a traditional perioperative analgesic method for its precise pain control and low systemic toxicity. However, a single low dose of local anesthetic merely provides a few hours of analgesia, and high dose results in irreversible toxicity, whereas continuous infusion of anesthetics is expensive and complicated. Therefore, it is necessary to develop a long-acting and sensory-selective local anesthetic for safe perioperative analgesia. Methods: An injectable composite comprising ropivacaine-loaded poly (ε-caprolactone) electrospun fiber and clonidine-loaded F127 hydrogel (Fiber-Rop/Gel-Clo composite) was developed for long-acting and walking regional analgesia with barely one dose. The peripheral nerve blockade effect of the composite was evaluated in a rat sciatic nerve block model. Also, the biodegradability and biosafety of the composite was evaluated. Results: The preferentially released Clo from the hydrogel rapidly constricted the peripheral arterial vessels, reducing the blood absorption of Rop and thus enhancing the local Rop accumulation at the injection site. The subsequently sustainable release of Rop from the fiber, significantly prolonged the sciatic nerve block of rats. Remarkably, an amazing sensorimotor segregation effect was achieved, as the sensory blockade (32.0 ± 1.4 h) lasted significantly longer than the motor blockade (20.3 ± 0.9 h). Additionally, the Fiber-Rop/Gel-Clo composite presented good biodegradability and biosafety in vivo. Conclusions: Our designed Fiber-Rop/Gel-Clo composite with minimal invasion, prolonged synergistic analgesia, and strikingly sensorimotor segregation effect, posted a promising prospect for regional long-term walking analgesia in clinical treatment.

A Multi-Omics Approach for Rapid Identification of Large Genomic Lesions at the Wheat Dense Spike (wds) Locus.

Optimal spike architecture provides a favorable structure for grain development and yield improvement. However, the number of genes cloned to underlie wheat spike architecture is extremely limited. Here, we obtained a wheat dense spike mutant (wds) induced by 60Co treatment of a common wheat landrace Huangfangzhu that exhibited significantly reduced spike and grain lengths. The shortened spike length was caused by longitudinal reduction in number and length of rachis cells. We adopted a multi-omics approach to identify the genomic locus underlying the wds mutant. We performed Exome Capture Sequencing (ECS) and identified two large deletion segments, named 6BL.1 at 334.8∼424.3 Mb and 6BL.2, 579.4∼717.8 Mb in the wds mutant. RNA-seq analysis confirmed that genes located in these regions lost their RNA expression. We then found that the 6BL.2 locus was overlapping with a known spike length QTL, qSL6B.2. Totally, 499 genes were located within the deleted region and two of them were found to be positively correlated with long spike accessions but not the ones with short spike. One of them, TraesCS6B01G334600, a well-matched homolog of the rice OsBUL1 gene that works in the Brassinosteroids (BR) pathway, was identified to be involved in cell size and number regulation. Further transcriptome analysis of young spikes showed that hormone-related genes were enriched among differentially expressed genes, supporting TraesCS6B01G334600 as a candidate gene. Our work provides a strategy to rapid locate genetic loci with large genomic lesions in wheat and useful resources for future wheat study.

InDels Identification and Association Analysis with Spike and Awn Length in Chinese Wheat Mini-Core Collection.

Diversity surveys of germplasm are important for gaining insight into the genomic basis for crop improvement; especially InDels, which are poorly understood in hexaploid common wheat. Here, we describe a map of 89,923 InDels from exome sequencing of 262 accessions of a Chinese wheat mini-core collection. Population structure analysis, principal component analysis and selective sweep analysis between landraces and cultivars were performed. Further genome-wide association study (GWAS) identified five QTL (Quantitative Trait Loci) that were associated with spike length, two of them, on chromosomes 2B and 6A, were detected in 10 phenotypic data sets. Assisted with RNA-seq data, we identified 14 and 21 genes, respectively that expressed in spike and rachis within the two QTL regions that can be further investigated for candidate genes discovery. Moreover, InDels were found to be associated with awn length on chromosomes 5A, 6B and 4A, which overlapped with previously reported genetic loci B1 (Tipped 1), B2 (Tipped 2) and Hd (Hooded). One of the genes TaAGL6 that was previously shown to affect floral organ development was found at the B2 locus to affect awn length development. Our study shows that trait-associated InDels may contribute to wheat improvement and may be valuable molecular markers for future wheat breeding.

Transcriptome Analysis of Developing Wheat Grains at Rapid Expanding Phase Reveals Dynamic Gene Expression Patterns.

Grain development, as a vital process in the crop's life cycle, is crucial for determining crop quality and yield. The wheat grain expanding phase is the early process involving the rapid morphological changes and initiation of grain filling. However, little is known about the molecular basis of grain development at this stage. Here, we provide a time-series transcriptome profile of developing wheat grain at 0, 2, 4, 6, 8, and 10 days after pollination of the wheat landrace Chinese Spring. A total of 26,892 differentially expressed genes, including 1468 transcription factors, were found between adjacent time points. Co-expression cluster analysis and Gene Ontology enrichment revealed dynamic expressions of cell division and starch biosynthesis related structural genes and transcription factors. Moreover, diverse, differential and drastically varied expression trends of the key genes related to hormone metabolism were identified. Furthermore, ~30% of triads showed unbalanced expression patterns enriching for genes in multiple pivotal metabolic pathways. Hormone metabolism related genes, such as YUC10 (YUCCA flavin-containing monooxygenase 10), AOS2 (allene oxide synthase 2), CYP90D2 (cytochrome P450 90D2), and CKX1 (cytokinin dehydrogenase 1), were dominantly contributed by A or D homoeologs of the triads. Our study provided a systematic picture of transcriptional regulation of wheat grains at the early grain expanding phase which should deepen our understanding of wheat grain development and help in wheat yield improvement.

Matrilineal empowers wheat pollen with haploid induction potency by triggering postmitosis reactive oxygen species activity.

Reactive oxygen species (ROS) play important roles during anther and pollen development. DNA damage may cause chromosome fragmentation that is considered to underlie chromosome elimination for haploid induction by matrilineal pollen, a key step in MATRILINEAL-based double haploid breeding technology. But when and how DNA damage occurs is unknown. We performed comparative studies of wheat pollens from the wild-type and the CRISPR/Cas9 edited matrilineal mutant (mMTL). Chemical assays detected a second wave of ROS in mMTL pollen at the three-nuclei-stage and subsequently, along with reduced antioxidant enzyme activities. RNA-seq analysis revealed disturbed expression of genes for fatty acid biosynthesis and ROS homoeostasis. Gas chromatography-mass spectrometry measurement identified abnormal fatty acid metabolism that may contribute to defective mMTL pollen walls as observed using electron microscopy, consistent with the function of MTL as a phospholipase. Moreover, DNA damage was identified using TdT-mediated dUTP nick-end labelling and quantified using comet assays. Velocity patterns showed that ROS increments preceded that of DNA damage over the course of pollen maturation. Our work hypothesises that mMTL-triggered later-stage-specific ROS causes DNA damage that may contribute to chromosome fragmentation and hence chromosome elimination during haploid induction. These findings may provide more ways to accelerate double haploid-based plant breeding.

Wheat breeding history reveals synergistic selection of pleiotropic genomic sites for plant architecture and grain yield.

Diversity surveys of crop germplasm are important for gaining insights into the genomic basis for plant architecture and grain yield improvement, which is still poorly understood in wheat. In this study, we exome sequenced 287 wheat accessions that were collected in the past 100 years. Population genetics analysis identified that 6.7% of the wheat genome falls within the selective sweeps between landraces and cultivars, which harbors the genes known for yield improvement. These regions were asymmetrically distributed on the A and B subgenomes with regulatory genes being favorably selected. Genome-wide association study (GWAS) identified genomic loci associated with traits for yield potential, and two underlying genes, TaARF12 encoding an auxin response factor and TaDEP1 encoding the G-protein γ-subunit, were located and characterized to pleiotropically regulate both plant height and grain weight. Elite single-nucleotide haplotypes with increased allele frequency in cultivars relative to the landraces were identified and found to have accumulated over the course of breeding. Interestingly, we found that TaARF12 and TaDEP1 function in epistasis with the classical plant height Rht-1 locus, leading to propose a "Green Revolution"-based working model for historical wheat breeding. Collectively, our study identifies selection signatures that fine-tune the gibberellin pathway during modern wheat breeding and provides a wealth of genomic diversity resources for the wheat research community.

Intertruncal versus classical approach to the ultrasound-guided supraclavicular brachial plexus block for upper extremity surgery: study protocol for a randomized non-inferiority trial.

BACKGROUND: Ultrasound-guided intertruncal approach (IA) to the supraclavicular block (SB) is recently proposed as a new approach for local anesthetic (LA) injection in terms of the classical approach (CA) at the level of the first rib. The CA-SB has been proven to result in satisfying sensorimotor block, but associate with a high risk of intraneural injection. The aim of this randomized non-inferiority study is to explore whether IA-SB can obtain similar block dynamics, as the CA-SB, but avoiding an intraneural injection during the whole nerve block procedure. METHODS: The total 122 patients undergoing elective upper extremity surgery will be randomly allocated to receive either an IA-SB or a CA-SB using a double-injection (DI) technique. In the IA-SB group, a portion of LA (15 mL) is injected accurately to the intertruncal plane between the middle and lower trunks under real-time ultrasound guidance; then, the remaining volume (10 mL) is carefully distributed to the other intertruncal plane between the upper and middle trunks. In the CA-SB group, the DI technique will be carried out as described in Tran's study. The primary outcome is the percentage of patients with a complete sensory blockade at 20 min with a predefined non-inferiority margin of - 5%. The secondary outcomes include the sensory-motor blockade of all 4 terminal nerves, onset times of the individual nerves within 30 min, block-related variables, and adverse events. DISCUSSION: The results will provide sensory-motor blockade-related parameters and safety of the ultrasound-guided intertruncal approach to the supraclavicular block, thereby promoting clinical practice. TRIAL REGISTRATION: Chinese Clinical Trial Registry ChiCTR2000040199 . Registered on 25 November 2020.

The wheat AGL6-like MADS-box gene is a master regulator for floral organ identity and a target for spikelet meristem development manipulation.

The AGAMOUS-LIKE6 (AGL6)-like genes are ancient MADS-box genes and are functionally studied in a few model plants. The knowledge of these genes in wheat remains limited. Here, by studying a 'double homoeolog mutant' of the AGL6 gene in tetraploid wheat, we showed that AGL6 was required for the development of all four whorls of floral organs with dosage-dependent effect on floret fertility. Yeast two-hybrid analyses detected interactions of AGL6 with all classes of MADS-box proteins in the ABCDE model for floral organ development. AGL6 was found to interact with several additional proteins, including the G protein ß and γ (DEP1) subunits. Analysis of the DEP1-B mutant showed a significant reduction in spikelet number per spike in tetraploid wheat, while overexpression of AGL6 in common wheat increased the spikelet number per spike and hence the grain number per spike. RNA-seq analysis identified the regulation of several meristem activity genes by AGL6, such as FUL2 and TaMADS55. Our work therefore extensively updated the wheat ABCDE model and proposed an alternative approach to improve wheat grain yield by manipulating the AGL6 gene.