Osa-miR1873 fine-tunes rice immunity against Magnaporthe oryzae and yield traits.

MicroRNAs (miRNAs) are known to fine-tune growth, development, and stress-induced responses. Osa-miR1873 is a rice-specific miRNA targeting LOC_Os05g01790. Here, we show that Osa-miR1873 fine-tunes rice immunity against Magnaporthe oryzae and yield traits via LOC_Os05g01790. Osa-miR1873 was significantly upregulated in a susceptible accession but downregulated in a resistance accession at 24 h post-inoculation (hpi) of M. oryzae. Overexpressing Osa-miR1873 enhanced susceptibility to M. oryzae and compromised induction of defense responses. In contrast, blocking Osa-miR1873 through target mimicry compromised susceptibility to M. oryzae and enhanced induction of defense responses. Altered expression of Osa-miR1873 also resulted in some defects in yield traits, including grain numbers and seed setting rate. Moreover, overexpression of the target gene LOC_Os05g01790 increased rice blast disease resistance but severely penalized growth and yield. Taken together, we demonstrate that Osa-miR1873 fine-tunes the rice immunity-growth trade-off via LOC_Os05g01790, and blocking Osa-miR1873 could improve blast disease resistance without significant yield penalty. Thus, the Osa-miR1873-LOC_Os05g01790 regulatory module is valuable in balancing yield traits and blast resistance.

Does training improve diagnostic accuracy and inter-rater agreement in applying the Berlin radiographic definition of acute respiratory distress syndrome? A multicenter prospective study.

BACKGROUND: Poor inter-rater reliability in chest radiograph interpretation has been reported in the context of acute respiratory distress syndrome (ARDS), although not for the Berlin definition of ARDS. We sought to examine the effect of training material on the accuracy and consistency of intensivists' chest radiograph interpretations for ARDS diagnosis. METHODS: We conducted a rater agreement study in which 286 intensivists (residents 41.3%, junior attending physicians 35.3%, and senior attending physician 23.4%) independently reviewed the same 12 chest radiographs developed by the ARDS Definition Task Force ("the panel") before and after training. Radiographic diagnoses by the panel were classified into the consistent (n = 4), equivocal (n = 4), and inconsistent (n = 4) categories and were used as a reference. The 1.5-hour training course attended by all 286 intensivists included introduction of the diagnostic rationale, and a subsequent in-depth discussion to reach consensus for all 12 radiographs. RESULTS: Overall diagnostic accuracy, which was defined as the percentage of chest radiographs that were interpreted correctly, improved but remained poor after training (42.0 ± 14.8% before training vs. 55.3 ± 23.4% after training, p < 0.001). Diagnostic sensitivity and specificity improved after training for all diagnostic categories (p < 0.001), with the exception of specificity for the equivocal category (p = 0.883). Diagnostic accuracy was higher for the consistent category than for the inconsistent and equivocal categories (p < 0.001). Comparisons of pre-training and post-training results revealed that inter-rater agreement was poor and did not improve after training, as assessed by overall agreement (0.450 ± 0.406 vs. 0.461 ± 0.575, p = 0.792), Fleiss's kappa (0.133 ± 0.575 vs. 0.178 ± 0.710, p = 0.405), and intraclass correlation coefficient (ICC; 0.219 vs. 0.276, p = 0.470). CONCLUSIONS: The radiographic diagnostic accuracy and inter-rater agreement were poor when the Berlin radiographic definition was used, and were not significantly improved by the training set of chest radiographs developed by the ARDS Definition Task Force. TRIAL REGISTRATION: The study was registered at ClinicalTrials.gov (registration number NCT01704066 ) on 6 October 2012.

Protective effect of ethyl pyruvate on pancreas injury in rats with severe acute pancreatitis.

BACKGROUND: Systemic inflammatory mediators have an important role in the development of acute pancreatitis. In this study, we investigated the effect of ethyl pyruvate (EP) on pancreas injury in rats with severe acute pancreatitis (SAP) and its possible mechanism. METHODS: We randomly allocated rats into the following three experimental groups: control and SAP- and EP-treated. Then, we recorded the mortality rate. We harvested tissue specimens for morphological studies, streptavidin-peroxidase immunohistochemistry examination, and Western blot analysis. We tested the levels of pancreatic tissue malondialdehyde and the activity of serum amylase, myeloperoxidase in the pancreas. In addition, we studied nuclear factor-κB (NF-κB) activation, tumor necrosis factor-α levels, and high mobility group box 1 protein expression levels in the pancreas. RESULTS: Treatment with EP after SAP was associated with a reduction in the severity of SAP and pancreas injury. Treatment with EP significantly decreased the expression of tumor necrosis factor-α and high mobility group box 1, and ameliorated malondialdehyde concentration and myeloperoxidase activity in the pancreas in SAP rats. Compared with the SAP group, treatment with EP significantly decreased the number of inflammatory cell infiltration, markedly inhibited pancreatic NF-κB DNA binding, and increased the survival rates. CONCLUSIONS: This study demonstrates that preventing the activation of NF-κB by EP ameliorates tissue injury associated with experimental murine acute pancreatitis. This result provides an important insight into the molecular biology of acute pancreatitis.

[Construction of stable human umbilical vein endothelial cells line expressing short hairpin RNA (shRNA) targeting high mobility group box-1 gene].

OBJECTIVE: To construct the short hairpin RNA (shRNA) targeting high mobility group box-1 (HMGB1) and culture the stable human umbilical vein endothelial cell (HUVEC) line expressing this shRNA. METHODS: Based on the HMGB1 gene sequence, shRNA was designed, synthesized and subcloned into the pRNA-u6.1/Neo vector, while negative controls were also established. Then the recombinant vector was transfected into HUVEC cell line and the cell was screened with G418 and assayed by using real time reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. RESULTS: Restriction endonuclease digestion test and sequencing verification showed that the recombinant pRNA-u6.1/Neo vector expressing this shRNA targeting HMGB1 was successfully constructed and the stable HUVEC cell line expressing this shRNA was developed. The real time RT-PCR and Western blotting was used to detect that recombinant plasmid in HUVEC cell effect on expression of HMGB1 was reduced. (mRNA: 0.4635 ± 0.0342 vs. 1.0340 ± 0.0352, protein: 0.4510 ± 0.0200 vs. 1.0210 ± 0.0110, both P<0.05). CONCLUSION: The recombinant pRNA-u6.1/Neo vector expressing shRNA targeting HMGB1 was successfully constructed and the stable HUVEC cell line expressing this shRNA was developed, and therefore allowed further investigation regarding the function of HMGB1 gene in the HUVEC cell line.

[Therapeutic effects of unfractionated heparin on lipopolysaccharide-activated matrix metalloproteinase-9 and tissue inhibitors of metalloproteinase-1 in endothelial cells].

OBJECTIVE: To investigate the expressions of matrix metalloproteinase-9 and tissue inhibitors of metalloproteinase-1 (TIMP-1) in injured endothelial cells induced by lipopolysaccharide (LPS) and the effects of unfractionated heparin (UFH) on the level of expressions. METHODS: The human pulmonary microvascular endothelial cells (HPMECs) were injured by LPS (10 µg/ml). In UFH pretreatment group, the cells were interfered with 0.1 U/ml or 1 U/ml UFH within 15 minutes before stimulus of LPS. In control group, the cells were cultured in equal volume of phosphate buffered saline (PBS). The RNA of the respectively cells were extracted at 2, 6, 12 hours after stimulus, and the expressions of MMP-9 mRNA and TIMP-1 mRNA were detected by real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS: Compared with control group, the expressions of MMP-9 mRNA and TIMP-1 mRNA were increased after stimulation of LPS, and peaked at 12 hours (MMP-9 mRNA: 4.26±0.81 vs. 1.00±0.46, TIMP-1 mRNA: 4.93±0.08 vs. 1.00±0.13, both P<0.05), the change in TIMP-1 was more significant. While as UFH pretreatment could significantly down-regulated the mRNA expressions of MMP-9 and TIMP-1 (UFH 0.1 U/ml group MMP-9 mRNA: 2.74±0.30, TIMP-1 mRNA: 2.96±0.13; UFH 1 U/ml group MMP-9 mRNA: 3.08±0.48, TIMP-1 mRNA: 2.93±0.27, all P<0.05). There were no significant differences in mRNA expressions between two UFH groups. CONCLUSIONS: The expressions of MMP-9 and TIMP-1 of HPMEC injured by LPS were obviously increased, UFH might attenuate the injury via inhibiting the expressions of MMP-9 and TIMP-1.

[Effects of unfractionated heparin on expression of metalloproteinase-2 and -9 in acute lung injury rats].

OBJECTIVE: To determine the activation of matrix metalloproteinase (MMP-2, MMP-9) in rats with acute lung injury (ALI) and the effects of unfractionated heparin (UFH) on the levels. METHODS: Eighteen male Wistar rats were divided into control group, ALI group and UFH group, with 6 rats in each group by means of random number table. ALI was induced by administering a bolus injection of lipopolysaccharide (LPS) via the caudal vein at a dose of 6 mg/kg. In UFH group the rats were treated intravenously with 100 U/kg of UFH 15 minutes before the injection of LPS. In control group, the rats were treated with the same volume of normal saline. Serum levels of MMP-2 and MMP-9 were measured by enzyme linked immunosorbent assay (ELISA) at 1, 3, 6 hours via femoral vein. Six hours after the injection of reagents, the rats were sacrificed and lung tissue samples were collected for mRNA analysis of the MMP-2 and MMP-9 by real-time fluorescence quantitate reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS: Compared with control group, the content of MMP-2 and MMP-9 in serum of ALI group were increased, and reached the peak at 6 hours (MMP-2: 2.86±0.40 µg/L vs. 1.21±0.24 µg/L, MMP-9: 2.54±0.29 µg/L vs. 1.15±0.34 µg/L, both P<0.01); they were down-regulated in UFH group at 6 hours (MMP-2: 1.92±0.31 µg/L vs. 2.86±0.40 µg/L, MMP-9: 1.82±0.26 µg/L vs. 2.54±0.29 µg/L, both P<0.05). Compared with control group, the mRNA expressions of MMP-2 and MMP-9 in the lung tissue of ALI group were increased at 6 hours (MMP-2 mRNA: 1.88±0.09 vs. 1.00±0.10, MMP-9 mRNA: 3.15±0.47 vs. 1.00±0.17, both P<0.01); they were down-regulated in UFH group (MMP-2 mRNA: 1.26±0.14 vs. 1.88±0.09, P<0.01; MMP-9 mRNA: 2.06±0.68 vs. 3.15±0.47, P<0.05), but still above the control group (both P<0.05). CONCLUSIONS: The present study demonstrated that the level of MMP-2 and MMP-9 increased in rats with ALI. UFH could exert protective effects by inhibiting expression of MMP-2 and MMP-9 in serum and lung tissue, in both mRNA and protein expression.

[Protective effects of heparin against increased permeability of endothelial cells induced by lipopolysaccharide].

OBJECTIVE: To evaluate the effects of heparin on the changes in permeability and cytoskeleton of cultured human umbilical vascular endothelial cells (HUVECs) induced by lipopolysaccharide (LPS). METHODS: HUVECs were cultured in vitro and randomly assigned to blank control group, LPS group, heparin group, and LPS + heparin group, n=8. Cell viability was determined by methyl thiazolyl tetrazolium (MTT) method. Endothelial permeability was measured with the Transwell chamber models. F-actin of cytoskeletons was assayed with fluorescein isothiocyanate (FITC)-phalloidine. RESULTS: Compared with the blank control group, 10 mg/L and 100 mg/L of LPS significantly inhibited cell viability (0.695±0.015, 0.476±0.030 vs. 0.860±0.053, P<0.05 and P<0.01). Heparin 100 kU/L also inhibited cell viability (0.675±0.030 vs. 0.840±0.023, P<0.05). Increase in permeability of endothelium was induced by 10 mg/L of LPS, and it peaked at 4 hours, there was significant difference compared with blank control group (5.882±0.101 vs. 4.489±0.015, P<0.05). At the same time, LPS led to the reorganization of F-actin cytoskeleton and the formation of stress fibers. LPS+heparin decreased the increase in permeability of endothelium at 2-12 hours (2 hours: 4.382±0.053 vs. 5.084±0.129, 4 hours: 4.528±0.044 vs. 5.882±0.101, 6 hours: 4.381±0.089 vs. 5.479±0.125, 12 hours: 4.447±0.054 vs. 4.719±0.080, all P<0.05), and attenuated the reorganization of F-actin and the formation of stress fibers. CONCLUSION: Heparin attenuates LPS-induced increase in permeability of HUVECs and alterations in F-actin organization, thus protects endothelial barrier.

Pretreatment of Rho kinase inhibitor inhibits systemic inflammation and prevents endotoxin-induced acute lung injury in mice.

BACKGROUND: Systemic inflammatory mediators play an important role in the development of sepsis. In this study, we analyzed the role of Rho kinase in the activation of immune response and acute lung injury in a mouse model of sepsis. METHODS: C57BL/6J mice were randomly divided into three groups: control, LPS, and LPS+fasudil. We used a mouse model of endotoxemia that consists of intraperitoneal injection of a high dose of LPS (30 mg/kg); a Rho kinase inhibitor, fasudil (10 mg/kg), dissolved in sterile saline (1 µL/g body weight) was applied by intraperitoneal injection at 18 and 1 h before injection of LPS (LPS+fasudil group). The control mice received vehicle sterile saline only. Blood was collected and lungs were harvested at 3 and/or 6 h for analysis. RESULTS: At 3 and 6 h, the increased TNF-α and IL-1ß levels in plasma and MPO activity in lung tissue by LPS could be significantly inhibited by fasudil. In addition, LPS-induced histologic changes in the lungs at 6 h could be effectively reversed by fasudil pretreatment. Furthermore, pretreatment of mice with fasudil inhibited LPS-induced increasing of TNF-α, IL-1ß mRNA expression (3 and 6 h) and AP-1/DNA binding activity (3 h) in blood cells. In survival studies, fasudil (10 mg/kg), which was administered 18 and 1 h before the application of LPS, conferred a protection against lethality induced by LPS (30 mg/kg). CONCLUSION: These results suggest that Rho kinase may play a role in the pathology of systemic inflammation during early phase of sepsis, and the potential mechanism of action may be partly through the adjustment of AP-1 pathway.

[The effect of factor VII activating protease treatment on proliferation and collagen synthesis of cultured human pulmonary fibroblasts].

OBJECTIVE: To examine the influence of factor VII-activating protease (FSAP) treatment on cell proliferation and collagen synthesis in human pulmonary fibroblasts (HPF). METHODS: Cultured HPF were treated with heparin (10 mg/L), platelet-derived growth factor-BB (PDGF-BB, 20 µg/L), FSAP (12 mg/L), aprotinin (10 mg/L) separately and in different mixture. The proliferation of the HPF was determined by 5-bromodeoxyuridine (BrDU) incorporation. The collagen III mRNA was determined by real-time polymerase chain reaction (PCR), and its protein expression, as well as the phosphorylation of p42 / p44 mitogen activated protein kinase (MAPK) were both determined using Western blotting. RESULTS: PDGF BB significantly (all P < 0.05) induced cell proliferation (1.23 ± 0.06 vs. 1.01 ± 0.01 without heparin, 1.24 ± 0.04 vs. 0.98 ± 0.01 with heparin), collagen III mRNA transcription (1.79 ± 0.02 vs. 1.00 ± 0.00 without aprotinin, 1.57 ± 0.01 vs. 1.00 ± 0.00 with aprotinin), and collagen III protein expression (0.54 ± 0.26 vs. 0.17 ± 0.05 without aprotinin, 0.59 ± 0.31 vs. 0.24 ± 0.15 with aprotinin) in HPF. PDGF BB also significantly (both P < 0.05) induced p42 / p44 MAPK phosphorylation in HPF (0.89 ± 0.24 vs. 0.51 ± 0.17 without aprotinin, 0.97 ± 0.41 vs. 0.37 ± 0.05 with aprotinin). In the presence of heparin FSAP significantly (all P < 0.05) inhibited the PDGF BB induced HPF proliferation (0.92 ± 0.23 vs 1.19 ± 0.11); collagen III mRNA transcription (0.61 ± 0.02 vs. 1.79 ± 0.02) and collagen III protein expression (0.15 ± 0.12 vs. 0.54 ± 0.26). FSAP also inhibited PDGF induced p42 / p44 MAPK phosphorylation in HPF (0.57 ± 0.16 vs. 0.89 ± 0.24) significantly (P < 0.05). The presence of aprotinin lead to a reversal of the inhibitory effect of FSAP on collagen III mRNA transcription (2.37 ± 0.21 vs.0.61 ± 0.02), collagen III protein expression (0.55 ± 0.24 vs. 0.15 ± 0.12) and p42 / p44 MAPK phosphorylation (0.86 ± 0.41 vs. 0.57 ± 0.16, all P < 0.05). CONCLUSION: FSAP could inhibit the PDGF-BB induced proliferation and collagen III synthesis in HPF in vitro through the suppression of p42 / p44 MAPK phosphorylation.

Facile synthesis of magnetic molybdenum disulfide@graphene nanocomposite with amphiphilic properties and its application in solid-phase extraction for a wide polarity of insecticides in wolfberry samples.

A novel magnetic molybdenum disulfide@graphene (Fe3O4/MoS2@G) nanocomposite with amphiphilic properties was prepared via a co-mixing solvothermal method. To demonstrate the feasibility of Fe3O4/MoS2@G as a sorbent during sample preparation, it was employed for the magnetic solid phase extraction (MSPE) of ten pyrethroids, three triazoles and two acaricide pyridaben and picoxystrobin in an emulsified aqueous solution. Dichloromethane was used as the extractant to form an emulsified aqueous solution. Subsequently, the Fe3O4/MoS2@G sorbent with amphiphilic properties was used to retrieve 15 wide polarity insecticides from dichloromethane via MSPE. The proposed method has the advantage of being applicable to different polar pesticides, strengthening the capacity of enrichment and purification of target analytes. The π-π interaction between the hydrophilic and hydrophobic moieties of Fe3O4/MoS2@G and the aromatic rings of target analytes were responsible for the efficient sorption. Thus, a reliable, convenient, and efficient method for the analysis of 15 insecticides with wide polarity in wolfberry samples was established by coupling Fe3O4/MoS2@G nanocomposite MSPE with gas chromatography-mass spectrometry (GC-MS) analysis. The obtained linearity of this method was in the range from 1 to 5000 ng mL-1 for 15 analytes, with determination coefficients (R2) ≥0.9907. The limit of detection (LOD) for 15 insecticides was in the range from 0.1 to 5.0 ng g-1. The recoveries of 15 insecticides from spiked wolfberry samples were in the range from 71.41% to 110.53%, and RSD was less than 14.8%.

Roles of RAGE/ROCK1 Pathway in HMGB1-Induced Early Changes in Barrier Permeability of Human Pulmonary Microvascular Endothelial Cell.

Background: High mobility group box 1 (HMGB1) causes microvascular endothelial cell barrier dysfunction during acute lung injury (ALI) in sepsis, but the mechanisms have not been well understood. We studied the roles of RAGE and Rho kinase 1 (ROCK1) in HMGB1-induced human pulmonary endothelial barrier disruption. Methods: In the present study, the recombinant human high mobility group box 1 (rhHMGB1) was used to stimulate human pulmonary microvascular endothelial cells (HPMECs). The endothelial cell (EC) barrier permeability was examined by detecting FITC-dextran flux. CCK-8 assay was used to detect cell viability under rhHMGB1 treatments. The expression of related molecules involved in RhoA/ROCK1 pathway, phosphorylation of myosin light chain (MLC), F-actin, VE-cadherin and ZO-1 of different treated groups were measured by pull-down assay, western blot and immunofluorescence. Furthermore, we studied the effects of Rho kinase inhibitor (Y-27632), ROCK1/2 siRNA, RAGE-specific blocker (FPS-ZM1) and RAGE siRNA on endothelial barrier properties to elucidate the related mechanisms. Results: In the present study, we demonstrated that rhHMGB1 induced EC barrier hyperpermeability in a dose-dependent and time-dependent manner by measuring FITC-dextran flux, a reflection of the loss of EC barrier integrity. Moreover, rhHMGB1 induced a dose-dependent and time-dependent increases in paracellular gap formation accompanied by the development of stress fiber rearrangement and disruption of VE-cadherin and ZO-1, a phenotypic change related to increased endothelial contractility and endothelial barrier permeability. Using inhibitors and siRNAs directed against RAGE and ROCK1/2, we systematically determined that RAGE mediated the rhHMGB1-induced stress fiber reorganization via RhoA/ROCK1 signaling activation and the subsequent MLC phosphorylation in ECs. Conclusion: HMGB1 is capable of disrupting the endothelial barrier integrity. This study demonstrates that HMGB1 activates RhoA/ROCK1 pathway via RAGE, which phosphorylates MLC inducing stress fiber formation at short time, and HMGB1/RAGE reduces AJ/TJ expression at long term independently of RhoA/ROCK1 signaling pathway.

Suppression of rice miR168 improves yield, flowering time and immunity.

MicroRNA168 (miR168) is a key miRNA that targets Argonaute1 (AGO1), a major component of the RNA-induced silencing complex1,2. Previously, we reported that miR168 expression was responsive to infection by Magnaporthe oryzae, the causal agent of rice blast disease3. However, how miR168 regulates immunity to rice blast and whether it affects rice development remains unclear. Here, we report our discovery that the suppression of miR168 by a target mimic (MIM168) not only improves grain yield and shortens flowering time in rice but also enhances immunity to M. oryzae. These results were validated through repeated tests in rice fields in the absence and presence of rice blast pressure. We found that the miR168-AGO1 module regulates miR535 to improve yield by increasing panicle number, miR164 to reduce flowering time, and miR1320 and miR164 to enhance immunity. Our discovery demonstrates that changes in a single miRNA enhance the expression of multiple agronomically important traits.

[The role of p38 mitogen-activated protein kinase/nuclear factor-ΚB transduction pathway on coagulation disorders due to endothelial injury induced by sepsis].

OBJECTIVE: To determine the activation status of p38 mitogen-activated protein kinase (p38MAPK)/nuclear factor-ΚB (NF-ΚB) in coagulation disorders due to endothelial injury induced by sepsis. METHODS: Human umbilical vein endothelial cells (HUVECs) were exposed to plasma obtained from 22 patients suffering from sepsis. Plasma was also obtained from 8 healthy individuals to serve as negative control, and tumor necrosis factor-α (TNF-α) was used as positive control. Phosphorylation and activity of p38MAPK and NF-ΚB were determined with enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunofluorescence assay. RESULTS: The level of TNF-α (ng/L) in sepsis plasma was significantly higher than that in healthy plasma (155.68±89.74 vs. 5.00±0.47, P <0.01). Compared with healthy plasma in 20% concentration it was found when HUVECs were treated with sepsis plasma in 20% concentration, tissue factor (TF, µg/L) reached the peak at 180 minutes (5.87±0.14 vs. 1.25±0.11, P <0.01), von Willebrand factor (vWF, µg/L) reached the peak at 120 minutes (9.59±0.07 vs. 3.59±0.06, P <0.01), then they began to decline. When HUVECs were treated with sepsis plasma in 20% concentration increased phosphorylation and activity of p38MAPK and NF-ΚB, phosphorylation of p38MAPK occurred before phosphorylation of NF-ΚB (2 minutes vs. 5 minutes). When the inhibitor of p38MAPK (SB239063) was added, NF-ΚB phosphorylation (activation) and NF-ΚB nuclear translocation were inhibited. CONCLUSION: This study demonstrates that p38MAPK/NF-ΚB transduction pathway plays an important role in septic coagulopathy.

Unfractionated heparin attenuates endothelial barrier dysfunction via the phosphatidylinositol-3 kinase/serine/threonine kinase/nuclear factor kappa-B pathway.

BACKGROUND: Vascular endothelial dysfunction is considered a key pathophysiologic process for the development of acute lung injury. In this study, we aimed at investigating the effects of unfractionated heparin (UFH) on the lipopolysaccharide (LPS)-induced changes of vascular endothelial-cadherin (VE-cadherin) and the potential underlying mechanisms. METHODS: Male C57BL/6 J mice were randomized into three groups: vehicle, LPS, and LPS + UFH groups. Intraperitoneal injection of 30 mg/kg LPS was used to induce sepsis. Mice in the LPS + UFH group received subcutaneous injection of 8 U UFH 0.5 h before LPS injection. The lung tissue of the mice was collected for assessing lung injury by measuring the lung wet/dry (W/D) weight ratio and observing histological changes. Human pulmonary microvascular endothelial cells (HPMECs) were cultured and used to analyze the effects of UFH on LPS- or tumor necrosis factor-alpha (TNF-α)-induced vascular hyperpermeability, membrane expression of VE-cadherin, p120-catenin, and phosphorylated myosin light chain (p-MLC), and F-actin remodeling, and on the LPS-induced activation of the phosphatidylinositol-3 kinase (PI3K)/serine/threonine kinase (Akt)/nuclear factor kappa-B (NF-κB) signaling pathway. RESULTS: In vivo, UFH pretreatment significantly attenuated LPS-induced pulmonary histopathological changes (neutrophil infiltration and erythrocyte effusion, alveolus pulmonis collapse, and thicker septum), decreased the lung W/D, and increased protein concentration (LPS vs. LPS + UFH: 0.57 ±â€Š0.04 vs. 0.32 ±â€Š0.04 mg/mL, P = 0.0092), total cell count (LPS vs. LPS + UFH: 9.57 ±â€Š1.23 vs. 3.65 ±â€Š0.78 × 10/mL, P = 0.0155), polymorphonuclear neutrophil percentage (LPS vs. LPS + UFH: 88.05% ±â€Š2.88% vs. 22.20% ±â€Š3.92%, P = 0.0002), and TNF-α (460.33 ±â€Š23.48 vs. 189.33 ±â€Š14.19 pg/mL, P = 0.0006) in the bronchoalveolar lavage fluid. In vitro, UFH pre-treatment prevented the LPS-induced decrease in the membrane expression of VE-cadherin (LPS vs. LPS + UFH: 0.368 ±â€Š0.044 vs. 0.716 ±â€Š0.064, P = 0.0114) and p120-catenin (LPS vs. LPS + UFH: 0.208 ±â€Š0.018 vs. 0.924 ±â€Š0.092, P = 0.0016), and the LPS-induced increase in the expression of p-MLC (LPS vs. LPS + UFH: 0.972 ±â€Š0.092 vs. 0.293 ±â€Š0.025, P = 0.0021). Furthermore, UFH attenuated LPS- and TNF-α-induced hyperpermeability of HPMECs (LPS vs. LPS + UFH: 8.90 ±â€Š0.66 vs. 15.84 ±â€Š1.09 Ω·cm, P = 0.0056; TNF-α vs. TNF-α + UFH: 11.28 ±â€Š0.64 vs. 18.15 ±â€Š0.98 Ω·cm, P = 0.0042) and F-actin remodeling (LPS vs. LPS + UFH: 56.25 ±â€Š1.51 vs. 39.70 ±â€Š1.98, P = 0.0027; TNF-α vs. TNF-α + UFH: 55.42 ±â€Š1.42 vs. 36.51 ±â€Š1.20, P = 0.0005) in vitro. Additionally, UFH decreased the phosphorylation of Akt (LPS vs. LPS + UFH: 0.977 ±â€Š0.081 vs. 0.466 ±â€Š0.035, P = 0.0045) and I kappa B Kinase (IKK) (LPS vs. LPS + UFH: 1.023 ±â€Š0.070 vs. 0.578 ±â€Š0.044, P = 0.0060), and the nuclear translocation of NF-κB (LPS vs. LPS + UFH: 1.003 ±â€Š0.077 vs. 0.503 ±â€Š0.065, P = 0.0078) in HPMECs, which was similar to the effect of the PI3K inhibitor, wortmannin. CONCLUSIONS: The protective effect of UFH against LPS-induced pulmonary endothelial barrier dysfunction involves VE-cadherin stabilization and PI3K/Akt/NF-κB signaling.

High salt-induced weakness of anti-oxidative function of natriuretic peptide receptor-C and podocyte damage in the kidneys of Dahl rats.

BACKGROUND: Atrial natriuretic peptide (ANP) and its natriuretic peptide receptors A (NPR-A) and C (NPR-C) are involved in the regulation of physiological and pathophysiological process of blood pressure. The present study aimed to determine the role of NPR-C in the development of salt-sensitive hypertension. METHODS: The Dahl salt-sensitive (DS) and salt-resistant (DR) rats were used in this study. Animals were matched according to their age and weight, and then placed on either a high-salt (HS, 8%) or a normal-salt (NS, 0.4%) diet for 6 weeks randomly using random number table. The systolic blood pressure (SBP), plasmatic sodium concentration (PLNa), urinary sodium excretion (UVNa), and serum creatinine concentration (Scr) were measured. The concentration of ANP in blood and tissues (heart and kidney) was detected by enzyme-linked immunosorbent assay. The expression of ANP, NPR-A, and NPR-C in kidney was evaluated with western blot analysis. Regarding renal redox state, the concentration changes in malondialdehyde (MDA), lipofuscin, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox), and nitric oxide synthase (NOS) in kidney were detected by a spectrophotometric method. The kidney damage was evaluated using pathological techniques and the succinodehydrogenase (SDHase) examination. Furthermore, after an intra-peritoneal injection of C-atrial natriuretic peptide (ANP)4-23 (C-ANP4-23), an NPR-C receptor agonist, the SBP, biochemical values in blood and urine, and renal redox state were evaluated. The paired Student's t test and analysis of variance followed by the Bonferroni test were performed for statistical analyses of the comparisons between two groups and multiple groups, respectively. RESULTS: The baseline SBP in all groups was within the normal range. At the end of the 6-week experiment, HS diet significantly increased the SBP in DS rats from 116.63 ±â€Š2.90 mmHg to 162.25 ±â€Š2.15 mmHg (t = -10.213, P < 0.001). The changes of SBP were not significant in DS rats on an NS diet and DR rats on an NS diet or on an HS diet (all P > 0.05). The significant increase of PLNa, UVNa, and Scr related to an HS diet was found in both DS and DR rats (all P < 0.05). However, significant changes in the concentration (t = -21.915, P < 0.001) and expression of renal ANP (t = -3.566, P = 0.016) and the expression of renal NPR-C (t = 5.864, P = 0.002) were only observed in DS hypertensive rats. The significantly higher desmin immunochemical staining score (t = -5.715, P = 0.005) and mitochondrial injury score (t = -6.325, P = 0.003) accompanied by the lower SDHase concentration (t = 3.972, P = 0.017) revealed mitochondrial pathologic abnormalities in podocytes in DS rats with an HS diet. The distinct increases of MDA (t = -4.685, P = 0.009), lipofuscin (t = -8.195, P = 0.001), and Nox (t = -12.733, P < 0.001) but not NOS (t = -0.328, P = 0.764) in kidneys were also found in DS hypertensive rats. C-ANP4-23 treatment significantly decreased the SBP induced by HS in DS rats (P < 0.05), which was still higher than NS groups with the vehicle or C-ANP4-23 treatment (P < 0.05). Moreover, the HS-induced increase of MDA, lipofuscin, Nox concentrations, and Nox4 expression in DS rats was significantly attenuated by C-ANP4-23 treatment as compared with those with HS diet and vehicle injection (all P < 0.05). CONCLUSIONS: The results indicated that the renal NPR-C might be involved in the salt-sensitive hypertension through the damage of mitochondria in podocytes and the reduction of the anti-oxidative function. Hence, C-ANP4-23 might serve as a therapeutic agent in treating salt-sensitive hypertension.

Osa-miR162a fine-tunes rice resistance to Magnaporthe oryzae and Yield.

MicroRNAs (miRNAs) play essential roles in rice immunity against Magnaporthe oryzae, the causative agent of rice blast disease. Here we demonstrate that Osa-miR162a fine-tunes rice immunity against M. oryzae and yield traits. Overexpression of Osa-miR162a enhances rice resistance to M. oryzae accompanying enhanced induction of defense-related genes and accumulation of hydrogen peroxide (H2O2). In contrast, blocking Osa-miR162 by overexpressing a target mimic of Osa-miR162a enhances susceptibility to blast fungus associating with compromised induction of defense-related gene expression and H2O2 accumulation. Moreover, the transgenic lines overexpressing Osa-miR162a display decreased seed setting rate resulting in slight reduced yield per plant, whereas the transgenic lines blocking Osa-miR162 show an increased number of grains per panicle, resulting in increased yield per plant. Altered accumulation of Osa-miR162 had a limited impact on the expression of rice Dicer-like 1 (OsDCL1) in these transgenic lines showing normal gross morphology, and silencing of OsDCL1 led to enhanced resistance to blast fungus similar to that caused by overexpression of Osa-miR162a, suggesting the involvement of OsDCL1 in Osa-miR162a-regulated resistance. Together, our results indicate that Osa-miR162a is involved in rice immunity against M. oryzae and fine-tunes resistance and yield.

Expressing a Target Mimic of miR156fhl-3p Enhances Rice Blast Disease Resistance Without Yield Penalty by Improving SPL14 Expression.

MicroRNAs (miRNAs) play essential roles in the regulation of plant growth and defense responses. More and more, miRNA-3ps are reported to act in plant development and immunity. miR156 is a conserved miRNA, and most previous studies focus on its roles in plant growth, development, and yield determinacy. Here, we show that expressing a target mimic of miR156fhl-3p led to enhanced rice blast disease resistance without a yield penalty. miR156fhl-3p was differentially responsive to Magnaporthe oryzae in susceptible and resistant accessions. Transgenic lines expressing a target mimic of miR156fhl-3p (MIM156-3p) exhibited enhanced rice blast disease resistance and increased expression of defense-related genes. MIM156-3p also enhanced the mRNA abundance of SPL14 and WRKY45 by down-regulating miR156-5p and pre-miR156. Moreover, MIM156-3p lines displayed a decreased number of second rachis branches per panicle but enlarged grains, leading to unchanged yield per plant. Consistently, overexpressing miR156h (OX156) led to enhanced susceptibility to M. oryzae and decreased the expression of SPL14 and WRKY45. Our results indicate that miR156fhl-3p mounts a regulatory role on miR156-5p, which subsequently regulates the expression of SPL14 and WRKY45 to improve rice blast disease resistance.

[Evaluation of clinical effects on low-dose heparin therapy for sepsis].

OBJECTIVE: To investigate the therapeutic effects of low-dose heparin on sepsis. METHODS: Seventy-nine sepsis patients were randomly divided into two groups: heparin treatment group (n = 37) and routine treatment group (n = 42). The 7-day and 28-day mortality, the days in ICU and the length of stay, the changes of oxygenation index, the days of mechanical ventilation and the rates of disseminated intravascular coagulation (DIC), acute renal failure (ARF), acute respiratory distress syndrome (ARDS) and multiple organ dysfunction syndrome (MODS) were observed. The levels of APTT, PT and platelet (PLT) count were determined before and after treatment in two groups. RESULTS: The rates of DIC, ARF and MODS in heparin group decreased significantly after therapy: rate of DIC, 15.4% vs 38.7% (P = 0.03); rate of ARF, 25.0% vs 51.9% (P = 0.04); rate of MODS, 26.3% vs 50.0% (P = 0.04). In heparin group, the 28-day mortality was statistically reduced (15.4% vs 32.4%, P = 0.03). The differences between heparin group and routine group were not statistically significant in the 7-day mortality (7.7% vs 12.9%, P = 0.08), the days in ICU (Z = 0.281, P = 0.779, rank sum test), the length of stay (Z = 0.562, P = 0.574, rank sum test), the oxygenation index (P = 0.82), the days of mechanical ventilation [(126.07 +/- 166.21) h vs (179.27 +/- 221.7) h, P = 0.28] and the rate of ARDS (44.0% vs 46.2%, P = 0.88). The differences in APTT, PT and PLT were not significant between the two groups. CONCLUSION: Low-dose heparin can decrease the mortality rate of sepsis and improve the prognosis of patients. It is a safe promising therapy in sepsis patients without severe side effects.

Prognostic value of red blood cell distribution width for severe acute pancreatitis.

BACKGROUND: Severe acute pancreatitis (SAP) is a common condition in the intensive care unit (ICU) and has a high mortality. Early evaluation of the severity and prognosis is very important for SAP therapy. Recently, red blood cell distribution (RDW) was associated with mortality of sepsis patients and could be used as a predictor of prognosis. Similarly, RDW may be associated with the prognosis of SAP patients and be used as a prognostic indicator for SAP patients. AIM: To investigate the prognostic value of RDW for SAP patients. METHODS: We retrospectively enrolled SAP patients admitted to the ICU of the First Affiliated Hospital of China Medical University from June 2015 to June 2017. According to the prognosis at 90 d, SAP patients were divided into a survival group and a non-survival group. RDW was extracted from a routine blood test. Demographic parameters and RDW were recorded and compared between the two groups. The receiver operator characteristic (ROC) curve was constructed and Cox regression analysis was performed to investigate the prognostic value of RDW for SAP patients. RESULTS: In this retrospective cohort study, 42 SAP patients were enrolled, of whom 22 survived (survival group) and 20 died (non-survival group). The baseline parameters were comparable between the two groups. The coefficient of variation of RDW (RDW-CV), standard deviation of RDW (RDW-SD), Acute Physiology and Chronic Health Evaluation II (APACHE II) score, and Sequential Organ Failure Assessment (SOFA) score were significantly higher in the non-survival group than in the survival group (P < 0.05). The RDW-CV and RDW-SD were significantly correlated with the APACHE II score and SOFA score, respectively. The areas under the ROC curves (AUCs) of RDW-CV and RDW-SD were all greater than those of the APACHE II score and SOFA score, among which, the AUC of RDW-SD was the greatest. The results demonstrated that RDW had better prognostic value for predicting the mortality of SAP patients. When the RDW-SD was greater than 45.5, the sensitivity for predicting prognosis was 77.8% and the specificity was 70.8%. Both RDW-CV and RDW-SD could be used as independent risk factors to predict the mortality of SAP patients in multivariate logistic regression analysis and univariate Cox proportional hazards regression analysis, similar to the APACHE II and SOFA scores. CONCLUSION: The RDW is greater in the non-surviving SAP patients than in the surviving patients. RDW is significantly correlated with the APACHE II and SOFA scores. RDW has better prognostic value for SAP patients than the APACHE II and SOFA scores and could easily be used by clinicians for the treatment of SAP patients.

Microbial effects on phosphorus release in aquatic sediments.

Microbial effects on phosphorus release were studied for the sediments of Tianjin source water by controlling DO and pH. The results show that: (1) In sterilised water, phosphorus began to release when pH = 9.1 and the stable release rate was 9.51 mg/(d.m2). It indicates that microorganisms may utilise anaerobic iron respiration to release Fe-P. (2) With unsterilised water, phosphorus release rate is 2.14 mg/(d.m2) when pH = 6.5, 8.60 mg/(d.m2) when pH is uncontrolled, and gets to 8.51 mg/(d.m2) when pH = 9.1. This indicates that microorganisms can dissolve insoluble phosphates to accelerate the ion exchange of OH(-) and PO4(3-), which are derived from iron-bound ortho-P and aluminium-bound ortho-P.