DataSheet_1_Case Report: Metagenomic next-generation sequencing assists in dynamic pathogen monitoring: powerful tool for progressing severe pneumonia.docx

BackgroundSevere community-acquired pneumonia (sCAP) is life-threatening and characterized by intensive care unit (ICU) admission and high mortality. And they are vulnerable to hospital-acquired infection. In such a severe condition, metagenomic next-generation sequencing (mNGS) outperforms for short turnaround time and broad detection spectrum.Case presentationA 15-year-old male with severe influenza and methicillin-resistant Staphylococcus aureus (MRSA) pneumonia progressed rapidly, initially misdiagnosed as influenza co-infected with Aspergillus for misleading bronchoscopy manifestations. The turnaround time of mNGS is 13 h, which has the potential to expedite the clinical medication process. With the powerful support of mNGS and extracorporeal membrane oxygenation (ECMO), anti-infective therapy was adjusted accordingly, and vital signs gradually stabilized. After tortuous treatment and unremitting efforts, the patient recovered well.ConclusionsRapid mNGS applications, timely medication adjustments, strong ECMO support and active family compliance contribute to this miracle of life. False-negative or false-positive results are alarming, anti-infective medications should be adjusted after a comprehensive review of physical status and other indicators.</p

Table_1_Visceral Adiposity Index Is a Measure of the Likelihood of Developing Depression Among Adults in the United States.docx

BackgroundDepression is a serious mental disorder often accompanied by emotional and physiological disorders. Visceral fat index (VAI) is the current standard method in the evaluation of visceral fat deposition. In this study, we explored the association between VAI and depression in the American population using NHANES data.MethodsA total of 2,577 patients were enrolled for this study. Data were collected through structured questionnaires. Subgroup analysis for the relationship between VAI and depression was evaluated using multivariate regression analysis after adjustment for potential confounding factors.ResultsFor every 1 unit increase in VAI, the clinical depression increased by 14% (OR = 1.14, 95% CI: 1.04–1.25). High VAI scores (T3) increased the highest risk of developing depression (OR = 2.32, 95% CI: 1.2–4.47). Subgroup analysis demonstrated a strong and stable association between VAI and the development of depression.ConclusionOur study showed that depressive symptoms are associated with a high ratio of visceral adiposity index after controlling confounding factors.</p

Suppression of the DIP:mDia2 axis impacts the CXCL12-driven blebbing mechanism.

<p>(A) MDA-MB-231 cells were transfected for 72 hrs with either GAPDH, DIP pool or individual DIP siRNAs and knockdown validated by i.p.-western blotting, with tubulin shown as a i.p. input control blot. (B) At 72 hrs post transfection, MDA-MB-231 cells were plated upon glass coverslips and stimulated for the indicated times with 30 ng/ml CXCL12 and blebbing cells enumerated, where n>100 cells. (C) HEK293T cells were transfected with 3XFLAG-mDia2 and either YFP, YFP-DIP or YFP-DIP N555A. Lysates were co-immunoprecipitated for DIP-associated mDia2. Tubulin is shown as a loading control for i.p. lysates. (D, E) MDA-MB-231 cells co-transfected for 48 hrs with 3XFLAG-mDia2 and YFP, YFP-DIP or YFP-DIP N555A were stimulated with 25 ng/ml CXCL12. Lysates were co-immunoprecipitated for DIP-associated mDia2 upon CXCL12 stimulation (D) and blebbing (E) was quantified as above. (F) MDA-MB-231 LN Luc cells stably expressing inducible miRNA directed against mDia2 were uninduced or induced with Doxycycline for 72 hrs and protein depletion validated (inset) by western blotting. (Un)induced cells were plated upon glass coverslips after 72 hrs induction and stimulated for the indicated times with 30 ng/ml CXCL12 and blebbing cells enumerated, where n>105 cells. (G) MDA-MB-231 cells were plated upon glass coverslips and, upon addition of either DMSO or 10 µm SMIFH2, blebbing was quantified in n>100 cells. Inset shows representative treated cells that were fixed and stained with phalloidin.</p

Additional file 1: of Genetic polymorphism in DGCR8 is associated with late onset of preeclampsia

Table S1. SNPs evaluated in this study and their minor allele frequencies (MAF) in the Han-Chinese population. (DOCX 17 kb

DIP LRR induces amoeboid motility in invading mesenchymal breast cancer cells.

<p>(A, B) MDA-MB-231 cells were treated with DMSO vehicle, 20 µm MMP-inhibitor GM6001 or were transfected with the indicated CFP-fusion proteins and were embedded in type-I collagen gels and stained as above. Insets validate the CFP-transfection. (C) Elongation indices were calculated as in 3C. (D) MDA-MB-231 cells transfected with either CFP-DIP LRR or CFP-DIP LRR N555A for 48 hrs were embedded in collagen gels overnight, stained with phalloidin and anti-pMLC2 antibodies. (E) MDA-MB-231 cells transfected for 48 hrs with CFP, CFP-DIP LRR or CFP-DIP LRR treated with 90 µM Y27632 reverse-invaded for 24 hrs through transwells coated with collagen I gels. Upon fixation, transwells were imaged by confocal microscopy. CFP pixels were calculated for 10 µm optical slices.</p

Image_2_Type 1 Cytotoxic T Cells Increase in Placenta after Intrauterine Inflammation.tif

CD8+ T cells recognize non-self antigen by MHC class I molecules and kill the target cells by the release of proinflammatory cytokines such as interferon gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α). Our group previously reported an increase of CD8+ T‐cell trafficking in the placenta with exposure to Lipopolysaccharides (LPS). CD8+ cytotoxic T cells have been classified into distinct subsets based upon cytokine production: Tc1 cells produce IFN-γ, Tc2 cells produce interleukin 4 (IL-4). Accordingly, the purpose of this research is to analyze the subsets of placenta CD8+ T cells. We hypothesized that LPS injection would induce a change of properties of CD8+ T cell and Tc1/Tc2 ratio. We investigated the subsets of CD8+ T cell infiltration to placenta and their specific function in response to LPS-induced inflammation in a mouse model. At embryonic (E) day 17, pregnant CD-1 dams received an intrauterine injection of 25 µg LPS in100 μl PBS or 100 μl of PBS only. Flow cytometry was used to quantify CD8+ T cells, evaluate the phenotype and subtypes, and detect markers of Tc1 and Tc2 cells in placenta, at 6 hours and 24 hours post injection (hpi). Intracellular staining and flow cytometry were performed to characterize cytokines produced by CD8+ T cells. Standard statistical analysis were employed. After 6 and 24 hours of LPS injection, total CD8 T cells increased (P<0.05). Tc1 cells expanded (P<0.05) in LPS-treated dams compared with the PBS group. The Tc1/Tc2 ratio was significantly higher in the LPS group than the PBS group (P<0.05). The expression of TNF-α and IFN-γ were increased in LPS group both at 6hpi and 24 hpi (P<0.05). We identified functional placental CD8+ T cell subtypes and found a significant increase ratio of Tc1/Tc2. Following IUI, CD8+ T cells induced inflammatory response in the placenta primarily via the production of Type 1 cytokines such as IFN-γ and TNF-α. We have provided evidence of a Tc1-bias response and cytokines in the mouse model of IUI.</p

CXCL12 induces an mDia2:DIP complex and membrane blebbing.

<p>(A) MDA-MB-231 cells were plated upon glass coverslips and stimulated for the indicated times with 30 ng/ml CXCL12. Blebbing cells were enumerated in shown triplicate experiments where n>54. Inset: representative blebs in a fixed cell stained with phalloidin. (B) Cells transfected with the designated constructs were stimulated with 25 ng/ml CXCL12. Lysates were co-immunoprecipitated for DIP-associated mDia2 upon CXCL12 stimulation. Tubulin is shown as an i.p input loading control. (C, D) Serum-starved MDA-MB-231 cells were adhered to glass coverslips and stimulated for 30 min with 30 ng/ml CXCL12 before fixation. Endogenous DIP, mDia2 and the F-actin architecture are shown using a 63x oil objective.</p

Dia-Interacting Protein (DIP) Imposes Migratory Plasticity in mDia2-Dependent Tumor Cells in Three-Dimensional Matrices

<div><p>Tumor cells rely upon membrane pliancy to escape primary lesions and invade secondary metastatic sites. This process relies upon localized assembly and disassembly cycles of F-actin that support and underlie the plasma membrane. Dynamic actin generates both spear-like and bleb structures respectively characterizing mesenchymal and amoeboid motility programs utilized by metastatic cells in three-dimensional matrices. The molecular mechanism and physiological trigger(s) driving membrane plasticity are poorly understood. mDia formins are F-actin assembly factors directing membrane pliancy in motile cells. mDia2 is functionally coupled with its binding partner DIP, regulating cortical actin and inducing membrane blebbing in amoeboid cells. Here we show that mDia2 and DIP co-tether to nascent blebs and this linkage is required for bleb formation. DIP controls mesenchymal/amoeboid cell interconvertability, while CXCL12 induces assembly of mDia2:DIP complexes to bleb cortices in 3D matrices. These results demonstrate how DIP-directed mDia2-dependent F-actin dynamics regulate morphological plasticity in motile cancer cells.</p> </div

Additional files 1: of Gestational weight gain in Chinese women -- results from a retrospective cohort in Changsha, China

Table S1. Subgroup analysis of comparison of adverse pregnancy outcomes between different gestational weight gain groups in the pre-pregnancy BMI 23.0–24.9 strata. (DOC 47 kb

Requirement for DIP in non-apoptotic membrane blebbing.

<p>(A–F) Constitutively blebbing M2 melanoma cells were plated upon glass coverslips for 3 hrs, fixed, permeabilized and stained simultaneously with phalloidin and antibodies directed against mDia2 or DIP. Cells are visualized using a 63x objective with a confocal microscope. (G) DIP (pool or individual siRNAs) or GAPDH (control siRNA) depletion in M2 cells was confirmed 72 hrs post-transfection by i.p.-western blotting. Tubulin was used as a loading control. (H) Upon 72 hrs of siRNA-mediated DIP (pool or individual) or GAPDH depletion, blebbing M2 cells were quantified. The experiment was performed in triplicate with n>50 cells per condition. (I) M2 cells treated with GAPDH, DIP pooled or individual siRNAs for 72 hrs were plated upon glass coverslips, stained for phalloidin and were visualized using a 63x objective by confocal microscopy.</p