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Agrobacterium-mediated transient expression (AMTE) has been widely used for high-throughput assays of gene function in diverse plant species. However, its application in monocots is still limited due to low expression efficiency. Here, by using histochemical staining and a quantitative fluorescence assay of β-glucuronidase (GUS) gene expression, we investigated factors affecting the efficiency of AMTE on intact barley plants. We found prominent variation in GUS expression levels across diverse vectors commonly used for stable transformation and that the vector pCBEP produced the highest expression. Additionally, concurrent treatments of plants with one day of high humidity and two days of darkness following agro-infiltration also significantly increased GUS expression efficiency. We thus established an optimized method for efficient AMTE on barley and further demonstrated its efficiency on wheat and rice plants. We showed that this approach could produce enough proteins suitable for split-luciferase assays of protein-protein interactions on barley leaves. Moreover, we incorporated the AMTE protocol into the functional dissection of a complex biological process such as plant disease. Based on our previous research, we used the pCBEP vector to construct a full-length cDNA library of genes upregulated during the early stage of rice blast disease. A subsequent screen of the library by AMTE identified 15 candidate genes (out of ~2000 clones) promoting blast disease on barley plants. Four identified genes encode chloroplast-related proteins: OsNYC3, OsNUDX21, OsMRS2-9, and OsAk2. These genes were induced during rice blast disease; however, constitutive overexpression of these genes conferred enhanced disease susceptibility to Colletotrichum higginsianum in Arabidopsis. These observations highlight the power of the optimized AMTE approach on monocots as an effective tool for facilitating functional assays of genes mediating complex processes such as plant-microbe interactions.

The evidence regarding the influence of allowing patients to participate in postoperative pain treatment decisions on acute pain management is contradictory. This study aimed to identify the role of patient participation in influencing pain-related patient-reported outcomes (PROs). This is a cross-sectional study. The data were provided by PAIN OUT (www.pain-out.eu). A dataset specific to adult Chinese patients undergoing orthopedic surgery was selected. The PROs were assessed on postoperative day 1. The patient participant was assessed using an 11-point scale. Participants who reported 5 were allocated to the "participation" group, and those who reported ≤5 were allocated to the "nonparticipation" group. A 1:1 propensity score matching was conducted. The primary outcome was the desire for more pain treatment. All other items of PROs were the secondary outcomes comprising pain intensity, interference of pain with function, emotional impairment, adverse effects, and other patient perception. From February 2014 to November 2020, 2244 patients from 20 centers were approached, of whom 1804 patients were eligible and 726 pairs were matched. There was no significant difference between the groups in the desire for more pain treatment either before (25.4% vs 28.2%, risk ratio [95% CI]: 0.90 [0.77, 1.05], P = .18) or after matching (26.7% vs 28.8%, risk ratio [95% CI]: 0.93 [0.79, 1.10], P = .43). After matching, patients in the participation group reported significantly better PROs, including pain intensity (less time spent in severe pain [P .01]), emotional impairment (less anxiety [P .01]), interference with function (less interference with sleep [P .01]), adverse effects (less drowsiness [P = .01]), and patient perception (more pain relief [P .01] and more satisfaction [P .01]), than the nonparticipation group. Patient participation in pain treatment decisions was associated with improved pain experience but failed to mitigate the desire for more treatment.

Objective: To screen out potential prognostic hub genes for adult patients with sepsis via construction of a microRNA-mRNA-PPI network, and investigate the localization of these core genes in peripheral blood monocytes.Methods: The peripheral bloods of 33 subjects were performed microRNA and mRNA sequencing using high-throughput sequencing, differentially expressed genes and microRNAs were identified by bioinformatics. 10×single-cell transcriptome sequencing was conducted further.Results: Of which 23 adult septic patients and 10 healthy individuals, a relative levels of transcription of 20391 genes, and 1633 microRNAs were detected by RNA sequencing. A total of 1114 differentially expressed genes and 76 differentially expressed microRNAs were obtained using DESeq2, 454 differential expression genes (DEGs) were distinguished ultimately. The microRNA-mRNA-PPI network was constructed based on DEGs and the top 20 differentially expressed microRNAs, which included 10 up-regulated and 10 down-regulated microRNAs. Furthermore, hub genes TLR5, FCGR1A, ELANE, GNLY, IL2RB and TGFBR3 which may be association with the prognosis of sepsis and their negative correlation microRNAs were recognized. Genes TLR5, FCGR1A and ELANE were mainly expressed in macrophages, genes GNLY, IL2RB and TGFBR3 were expressed specifically in T cells and nature kill cells.Conclusions: The parallel analysis of mRNA and microRNA in patients with sepsis using RNA-seq was feasible. Potential hub genes and microRNAs which are possibly related to sepsis prognosis were identified, it provides new prospects for sepsis. However, further experiments need to be required.

Hip fracture is a serious event in the older population and is associated with morbidity, mortality, and disability among those who survive. Emerging evidence suggests that frailty is pertinent to the clinical outcomes of older patients with hip fracture.We conducted a retrospective cohort study on older patients (aged ≥65 years) who underwent orthopedic repair with hip fracture under spinal or general anesthesia between June 1, 2019 and May 31, 2020. Demographic, surgical, and anesthetic features; chart-derived frailty index (CFI); occurrence of complications; and length of stay were retrospectively collected by reviewing patients' anesthesia records and medical charts. Patients were divided into 2 groups according to their CFI: high CFI group (CFI, 3-5) and low CFI group (CFI, 0-2). The CFI was calculated as the addition of the patient's conditions, including age70 years, preoperative body mass index18.5 kg/m2, hematocrit35%, albumin34 g/L, and serum creatinine176.8 mol/L (2.0 mg/dL). The EuroQol 5-dimensional questionnaire (EQ-5D) was obtained through telephone interviews 12 months after surgery to assess mortality and long-term quality of life.During the study period, 381 patients met the inclusion criteria. Patients in the high CFI group had an increased incidence of delirium by 13.80% (95% confidence interval [CI], 6.31-21.29) (17 [16.6%] vs 8 [2.8%]; P.001) compared to patients in the low CFI group. More patients in the high CFI group had pneumonia with an increased incidence of 17.71% (95% CI, 7.08-23.34) (40 [39.2%] vs 60 [21.5%]; P.001) than patients in the low CFI group. Postoperative hospital stay was significantly longer in the high CFI group (8 [6-12] vs 7 [5-10] days; P = .0222) than in the low CFI group. More patients died in the high CFI group 1 year after surgery with an increased mortality of 19.33% (95% CI, 9.47-29.18) (26.4% [23/87] vs 7.1% [16/225]; P.001) than patients in the low CFI group. A total of 64 patients in the high CFI group and 209 patients in the low CFI group completed the EQ-5D survey 1 year after surgery. The EQ-5D score in the high CFI group was significantly lower than that in the low CFI group (0.63 ± 0.22 vs 0.72 ± 0.22; P = .002).Frailty status is associated with adverse postoperative outcomes, mortality, and low quality of life 12 months after hip fracture surgery in older patients.