[Different methods in predicting mortality of pediatric intensive care units sepsis in Southwest China].

Objective: To investigate the value of systemic inflammatory response syndrome (SIRS), pediatric sequential organ failure assessment (pSOFA) and pediatric critical illness score (PCIS) in predicting mortality of pediatric sepsis in pediatric intensive care units (PICU) from Southwest China. Methods: This was a prospective multicenter observational study. A total of 447 children with sepsis admitted to 12 PICU in Southwest China from April 2022 to March 2023 were enrolled. Based on the prognosis, the patients were divided into survival group and non-survival group. The physiological parameters of SIRS, pSOFA and PCIS were recorded and scored within 24 h after PICU admission. The general clinical data and some laboratory results were recorded. The area under the curve (AUC) of the receiver operating characteristic curve was used to compare the predictive value of SIRS, pSOFA and PCIS in mortality of pediatric sepsis. Results: Amongst 447 children with sepsis, 260 patients were male and 187 patients were female, aged 2.5 (0.8, 7.0) years, 405 patients were in the survival group and 42 patients were in the non-survival group. 418 patients (93.5%) met the criteria of SIRS, and 440 patients (98.4%) met the criteria of pSOFA≥2. There was no significant difference in the number of items meeting the SIRS criteria between the survival group and the non-survival group (3(2, 4) vs. 3(3, 4) points, Z=1.30, P=0.192). The pSOFA score of the non-survival group was significantly higher than that of the survival group (9(6, 12) vs. 4(3, 7) points, Z=6.56, P<0.001), and the PCIS score was significantly lower than that of the survival group (72(68, 81) vs. 82(76, 88) points, Z=5.90, P<0.001). The predictive value of pSOFA (AUC=0.82) and PCIS (AUC=0.78) for sepsis mortality was significantly higher than that of SIRS (AUC=0.56) (Z=6.59, 4.23, both P<0.001). There was no significant difference between pSOFA and PCIS (Z=1.35, P=0.176). Platelet count, procalcitonin, lactic acid, albumin, creatinine, total bilirubin, activated partial thromboplastin time, prothrombin time and international normalized ratio were all able to predict mortality of sepsis to a certain degree (AUC=0.64, 0.68, 0.80, 0.64, 0.68, 0.60, 0.77, 0.75, 0.76, all P<0.05). Conclusion: Compared with SIRS, both pSOFA and PCIS had better predictive value in the mortality of pediatric sepsis in PICU.

Fluxes of Atmospheric Greenhouse-Gases in Maryland (FLAGG-MD): Emissions of Carbon Dioxide in the Baltimore, MD-Washington, D.C. area.

To study emissions of CO2 in the Baltimore, MD-Washington, D.C. (Balt-Wash) area, an aircraft campaign was conducted in February 2015, as part of the FLAGG-MD (Fluxes of Atmospheric Greenhouse-Gases in Maryland) project. During the campaign, elevated mole fractions of CO2 were observed downwind of the urban center and local power plants. Upwind flight data and HYSPLIT (Hybrid Single Particle Lagrangian Integrated Trajectory) model analyses help account for the impact of emissions outside the Balt-Wash area. The accuracy, precision, and sensitivity of CO2 emissions estimates based on the mass balance approach were assessed for both power plants and cities. Our estimates of CO2 emissions from two local power plants agree well with their CEMS (Continuous Emissions Monitoring Systems) records. For the 16 power plant plumes captured by the aircraft, the mean percentage difference of CO2 emissions was -0.3 %. For the Balt-Wash area as a whole, the 1σ CO2 emission rate uncertainty for any individual aircraft-based mass balance approach experiment was ±38 %. Treating the mass balance experiments, which were repeated seven times within nine days, as individual quantifications of the Balt-Wash CO2 emissions, the estimation uncertainty was ±16 % (standard error of the mean at 95% CL). Our aircraft-based estimate was compared to various bottom-up fossil fuel CO2 (FFCO2) emission inventories. Based on the FLAGG-MD aircraft observations, we estimate 1.9±0.3 MtC of FFCO2 from the Balt-Wash area during the month of February 2015. The mean estimate of FFCO2 from the four bottom-up models was 2.2±0.3 MtC.

Inhibition of hepatitis B virus replication in 2.2.15 cells by expressed shRNA.

Hepatitis B virus (HBV) infection is a worldwide health problem. To determine whether RNA interference (RNAi) could inhibit ongoing HBV replication in 2.2.15 cells, we constructed shRNA-producing vector pU6P based on the mouse U6 RNA promoter and cloned 12 targeted sequences against HBV into the vector, resulting in a series of pU6-siHBV vectors. The recombinant vectors were transfected into 2.2.15 cells, HBsAg and HBeAg in cultured media were assayed using enzyme-linked immunosorbent assay at various days after transfection. The amount of HBV DNA in the culture medium was quantitated by real-time polymerase chain reaction. HBsAg and HBeAg expression were inhibited by 72.8 +/- 5.4% (P = 0.00003) and 55.8 +/- 6.2% (P = 0.000026), respectively, 4 days after transfection with pU6-siHBV5. The greatest inhibition of HBV DNA was decreased by approximately 1.9-fold (P = 0.013) on day 6 post transfection with pU6-siHBV11 compared with that of empty vector. No change was found for HBV protein expression and DNA replication on pU6-siGFP (negative control) transfected cells. Our data demonstrate that the transfection of HBV-targeted shRNA-producing vector in 2.2.15 cells could inhibit the HBV protein expression and HBV DNA replication specifically. RNAi may be considered as a potential antiviral approach for human HBV infection.

Keratin 17 mutation in pachyonychia congenita type 2 with early onset sebaceous cysts.

BACKGROUND: Pachyonychia congenita (PC) is a group of autosomal dominant ectodermal dysplasias caused by mutations in four differentiation-specific keratin genes. Two major clinical subtypes of PC have been generally recognized. Symmetrically thickened fingernails and toenails are the defining characteristic of PC type 2 (PC-2) with onset at infancy. Pilosebaceous cysts are the best hallmark of PC-2, but they usually occur at puberty. OBJECTIVES: To report a Chinese pedigree of PC-2 with unusually early onset sebaceous cysts and to explore the genetic mutation and its phenotype. METHODS: Exon 1 of keratin 17 was amplified by polymerase chain reaction (PCR) from genomic DNA from the three patients in the pedigree, the proband, his half-sister and his younger son, two unaffected members in the pedigree and 50 unrelated and unaffected people. PCR products were directly sequenced to detect the mutation. RESULTS: Direct sequencing of the PCR products revealed a heterozygous 275A-->G mutation in all three affected members. This mutation predicts the substitution of asparagine by serine in codon 92 (N92S) located in the 1A domain of keratin 17. CONCLUSIONS: Mutation in the 1A domain of keratin 17 underlies the affected members' phenotype, PC-2 with early onset sebaceous cysts and late-onset thickened fingernails and toenails. The onset of the cysts is very early in some people within this family and the age at onset of thickened fingernails and toenails is variable within the family, implying the existence of modifying factors.

Inhibition of PYK2-induced actin cytoskeleton reorganization, PYK2 autophosphorylation and focal adhesion targeting by FAK.

Focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (PYK2) are structurally related tyrosine kinases. They are implicated in regulating actin cytoskeleton organization, a process critical for cell migration, mitosis and tumor metastasis. In this paper, we demonstrate that, although both PYK2 and FAK were expressed and colocalized at focal adhesions in fibroblasts, microinjection of PYK2, but not FAK, in Swiss 3T3 fibroblastic cells led to reorganization of focal adhesions and cell rounding. PYK2-mediated actin cytoskeleton reorganization required the PYK2 N terminus, the focal adhesion targeting (FAT) domain, catalytic activity and autophosphorylation. Remarkably, FAK suppressed PYK2-mediated reorganization of focal adhesions and cell rounding. In addition, FAK inhibited PYK2 autophosphorylation and focal adhesion targeting, which might contribute to FAK-mediated suppression of PYK2's phenotypes. Further analyses demonstrated that the inhibition of PYK2 autophosphorylation required the FAK N terminus but not FAK tyrosine phosphorylation. The FAK FAT domain seemed to be critical for FAK-mediated suppression of PYK2 focal adhesion targeting. Taken together, these results demonstrate that FAK could inhibit PYK2 autophosphorylation, focal adhesion targeting and actin cytoskeleton reorganization, suggesting that the balance between FAK and PYK2 tyrosine kinases is important for regulating cellular morphology, cell migration and cell growth.

Signal transduction in neuronal migration: roles of GTPase activating proteins and the small GTPase Cdc42 in the Slit-Robo pathway.

The Slit protein guides neuronal and leukocyte migration through the transmembrane receptor Roundabout (Robo). We report here that the intracellular domain of Robo interacts with a novel family of Rho GTPase activating proteins (GAPs). Two of the Slit-Robo GAPs (srGAPs) are expressed in regions responsive to Slit. Slit increased srGAP1-Robo1 interaction and inactivated Cdc42. A dominant negative srGAP1 blocked Slit inactivation of Cdc42 and Slit repulsion of migratory cells from the anterior subventricular zone (SVZa) of the forebrain. A constitutively active Cdc42 blocked the repulsive effect of Slit. These results have demonstrated important roles for GAPs and Cdc42 in neuronal migration. We propose a signal transduction pathway from the extracellular guidance cue to intracellular actin polymerization.

Regulation of CDC42 GTPase by proline-rich tyrosine kinase 2 interacting with PSGAP, a novel pleckstrin homology and Src homology 3 domain containing rhoGAP protein.

Proline-rich tyrosine kinase 2 (PYK2), a tyrosine kinase structurally related to focal adhesion kinase (FAK), is implicated in regulating cytoskeletal organization. However, mechanisms by which PYK2 participates in and regulates cytoskeletal organization remain largely unknown. Here we report identification of PSGAP, a novel protein that interacts with PYK2 and FAK and contains multiple domains including a pleckstrin homology domain, a rhoGTPase-activating protein domain, and a Src homology 3 domain. PYK2 interacts with PSGAP Src homology 3 domain via the carboxyl-terminal proline-rich sequence. PSGAP is able to increase GTPase activity of CDC42 and RhoA in vitro and in vivo. Remarkably, PYK2, but not FAK, can activate CDC42 via inhibition of PSGAP-mediated GTP hydrolysis of CDC42. Moreover, PSGAP is localized at cell periphery in fibroblasts in a pleckstrin homology domain-dependent manner. Over expression of PSGAP in fibroblasts results in reorganization of cytoskeletal structures and changes of cellular morphology, which requires rhoGTPase-activating activity. Taken together, our results suggest that PSGAP is a signaling protein essential for PYK2 regulation of cytoskeletal organization via Rho family GTPases.

[Determination of salicylic acid and its hydroxylated products using high performance liquid chromatography and fluorescence detection].

High performance liquid chromatography (HPLC) with fluorescence detector was used to determine salicylic acid (SA) and its main hydroxylated products, 2,3-dihydroxybenzoic acid (2,3-DHBA) and 2,5-dihydroxybenzoic acid (2,5-DHBA). Mobile phase was 30 mmol/L acetate(pH 4.9) and the flow rate was 1.0 mL/min. Low detection limits, 10 nmol/L for 2,3-DHBA, 0.6 nmol/L for 2,5-DHBA and 1.0 nmol/L for SA were obtained. The photolysis system of salicylic acid and H2O2 was investigated. The products in the system were measured and the yields were 45% and 51% for 2,3-DHBA and 2,5-DHBA, respectively.