Association between Fibrinogen-to-Albumin Ratio and Prognosis in Patients Admitted to an Intensive Care Unit.

The objective of this study was to investigate the usefulness of fibrinogen-to-albumin ratio (FAR) as a prognostic marker in patients admitted to an intensive care unit (ICU) compared with Sequential Organ Failure Assessment (SOFA) score, a widely used prognostic scoring system. An inverse probability weighting (IPW) was used to control for selection bias and confounding factors. After IPW adjustment, the high FAR group showed significantly higher risk of 1-year compared with low FAR group (36.4% vs. 12.4%, adjust hazard ratio = 1.72; 95% confidence interval (CI): 1.59-1.86; p < 0.001). In the receiver-operating characteristic curve analysis associated with the prediction of 1-year mortality, there was no significant difference between the area under the curve of FAR on ICU admission (C-statistic: 0.684, 95% CI: 0.673-0.694) and that of SOFA score on ICU admission (C-statistic: 0.679, 95% CI: 0.669-0.688) (p = 0.532). In this study, FAR and SOFA score at ICU admission were associated with 1-year mortality in patients admitted to an ICU. Especially, FAR was easier to obtain in critically ill patients than SOFA score. Therefore, FAR is feasible and might help predict long-term mortality in these patients.

The Effects of Body Composition, Physical Fitness on Time of Useful Consciousness in Hypobaric Hypoxia.

INTRODUCTION: Several previous studies have reported that hypoxia accidents of fighter pilots are rarer than gravity-induced loss of consciousness and spatial disorientation; however, the risk is greater. Therefore, this study aimed to investigate the relationship between physical fitness and body composition on time of useful consciousness (TUC) in hypobaric hypoxia. MATERIALS AND METHODS: Body composition and physical fitness testing on human participants were performed; subsequently, they were exposed to hypobaric hypoxia at a simulated altitude of 25,000 ft. Cognitive testing of the participants was accomplished by having them perform arithmetic task tables until they stopped writing for a period exceeding 5 seconds, at which point, they were placed on 100% oxygen. TUC was measured from the time the participants removed their oxygen masks to the time when the oxygen masks were placed back on them. Pearson's correlation was used to determine the relationship between TUC and other variables, and multiple regression was performed to determine the independent variables that best explain the TUC. RESULTS: TUC was positively correlated with the maximum oxygen uptake, stroke volume, arteriovenous oxygen difference, and endurance (sit-up and push-up). The maximum heart rate on the ground, high altitude, body fat mass, and percent body fat were negatively correlated with TUC. A regression analysis showed that 84.5% of the TUC can be explained by body composition and physical fitness. CONCLUSIONS: Our results revealed that increased cardiorespiratory fitness and decreased body fat mass could significantly impact the TUC. Therefore, for Air Force pilots who are frequently at high altitudes and at risk for exposure to hypoxia, aerobic exercise is significant to hypoxia tolerance.

Fabrication of Carbon Nanomaterials Using Laser Scribing on Copper Nanoparticles-Embedded Polyacrylonitrile Films and Their Application in a Gas Sensor.

Carbon nanomaterials have attracted significant research attention as core materials in various industrial sectors owing to their excellent physicochemical properties. However, because the preparation of carbon materials is generally accompanied by high-temperature heat treatment, it has disadvantages in terms of cost and process. In this study, highly sensitive carbon nanomaterials were synthesized using a local laser scribing method from a copper-embedded polyacrylonitrile (CuPAN) composite film with a short processing time and low cost. The spin-coated CuPAN was converted into a carbonization precursor through stabilization and then patterned into a carbon nanomaterial of the desired shape using a pulsed laser. In particular, the stabilization process was essential in laser-induced carbonization, and the addition of copper promoted this effect as a catalyst. The synthesized material had a porous 3D structure that was easy to detect gas, and the resistance responses were detected as -2.41 and +0.97% by exposure to NO2 and NH3, respectively. In addition, the fabricated gas sensor consists of carbon materials and quartz with excellent thermal stability; therefore, it is expected to operate as a gas sensor even in extreme environments.

Long-term exposures to low doses of silver nanoparticles enhanced in vitro malignant cell transformation in non-tumorigenic BEAS-2B cells.

To predict carcinogenic potential of AgNPs on the respiratory system, BEAS-2B cells (human bronchial epithelial cells) were chronically exposed to low- and non-cytotoxic dose (0.13 and 1.33µg/ml) of AgNPs for 4months (#40 passages). To assess malignant cell transformation of chronic exposure to AgNPs, several bioassays including anchorage independent agar colony formation, cell migration/invasion assay, and epithelial-mesenchymal transition (EMT) were performed in BEAS-2B cells. Chronic exposure to AgNPs showed a significant increase of anchorage independent agar colony formation and cell migration/invasion. EMT, which is the loss of epithelial markers (E-Cadherin and Keratin) and the gain of mesenchymal marker (N-cadherin and Vimentin), was induced by chronic exposure to AgNPs. These responses indicated that chronic exposure to AgNPs could acquire characteristics of tumorigenic cells from normal BEAS-2B cells. In addition, caspase-3, p-p53, p-p38, and p-JNK were significantly decreased, while p-ERK1/2 was significantly increased. MMP-9 related to cell migration/invasion was upregulated, while a MMP-9 inhibitor, TIMP-1 was down-regulated. These results indicated that BEAS-2B cells exposed to AgNPs could induce anti-apoptotic response/anoikis resistance, and cell migration/invasion by complex regulation of MAPK kinase (p38, JNK, and ERK) and p53 signaling pathways. Therefore, we suggested that long-term exposure to low-dose of AgNPs could enhance malignant cell transformation in non-tumorigenic BEAS-2B cells. Our findings provide useful information needed to assess the carcinogenic potential of AgNPs.

Small molecule-mediated duplex formation of nucleic acids with 'incompatible' backbones.

Proflavine, a known intercalator of DNA and RNA, promotes duplex formation by nucleic acids with natural and non-natural backbones that otherwise form duplexes with low thermal stability, and even some that show no sign of duplex formation in the absence of proflavine. These findings demonstrate the potential for intercalators to be used as cofactors for the assembly of rationally designed nucleic acid structures, and could provide fundamental insights regarding intercalation of natural nucleic acid duplexes.

Synthesis of phosphoramidites of isoGNA, an isomer of glycerol nucleic acid.

IsoGNA, an isomer of glycerol nucleic acid GNA, is a flexible (acyclic) nucleic acid with bases directly attached to its linear backbone. IsoGNA exhibits (limited) base-pairing properties which are unique compared to other known flexible nucleic acids. Herein, we report on the details of the preparation of isoGNA phosphoramidites and an alternative route for the synthesis of the adenine derivative. The synthetic improvements described here enable an easy access to isoGNA and allows for the further exploration of this structural unit in oligonucleotide chemistry thereby spurring investigations of its usefulness and applicability.

Textural interface opacity after Descemet-stripping automated endothelial keratoplasty.

PURPOSE: To describe cases of interface haze, also known as textural interface opacity, after Descemet-stripping automated endothelial keratoplasty (DSAEK). SETTING: Department of Cornea, External Disease, and Refractive Surgery Service, Duke Eye Center, Durham, North Carolina, USA. DESIGN: Retrospective case series. METHODS: Patients' clinical and demographic characteristics, cornea donor information, surgical technique, histopathology, and anterior segment optical coherence tomography (OCT) were reviewed retrospectively and clinical outcomes reported. RESULTS: The interface haze that developed after DSAEK comprised 2 types of textural interface opacity; that is, total (11 cases) and partial (3 cases). The time of onset of textural interface opacity ranged from 1 day to 7 weeks postoperatively. Although most patients with textural interface opacity showed improvement, with a corrected distance visual acuity better than 20/50, 3 had persistent decreased visual acuity and required repeat DSAEK. Seven eyes had concurrent phacoemulsification with intraocular lens implantation and DSAEK. The corneal graft was inserted with an Endoserter device in 11 eyes, an insertion forceps in 2 eyes, and a cystotome needle in 1 eye. Histopathology of the grafts of eyes that required repeat DSAEK showed no inflammation, no foreign-body deposit, and no fibrosis. Anterior segment OCT showed increased hyperreflectivity in the interface. CONCLUSIONS: Although the etiology of textural interface opacity is unclear, it may be related to retained ophthalmic viscosurgical device (OVD) or an adhesive property of the OVD used during the surgery. Although most cases resolve with time, repeat DSAEK may be an effective treatment for refractory cases. FINANCIAL DISCLOSURE: Dr. Kim is a consultant to Ocular Systems, Inc. No other author has a financial or proprietary interest in any material or method mentioned.

Asymmetric induction in 8π electrocyclizations. Design of a removable chiral auxiliary.

The pseudo C(2) symmetric trans diphenyl oxazoline group acts as an effective chiral auxiliary in the 8π, 6π tandem electrocyclization of a substituted tetraene 1-carboxylic acid. Assignment of absolute stereochemistry to the [4.2.0] bicyclooctadiene product supports a model in which both s-cis and s-trans conformations favor the transition states with the same helical twist. This assignment prefaces the development of analogs of SNF4435 C and D. These natural products demonstrate activity as androgen receptor antagonists and as multidrug resistance (mdr) reversal agents.

Construction of a humanized antibody to hepatitis B surface antigen by specificity-determining residues (SDR)-grafting and de-immunization.

We previously constructed a humanized antibody, HuS10, by grafting the complementarity-determining regions (CDRs) of a parental murine monoclonal antibody into the homologous human antibody sequences. This process is termed CDR grafting. Some residues that were thought to affect the CDR loops and stabilize the structure of the variable regions were retained in the framework region. HuS10 exhibited in vivo virus-neutralizing activity, but its murine content had the potential to elicit immune responses in patients. In this study, to minimize the immunogenic potential of HuS10, we replaced 17 mouse residues in HuS10 with the comparable human residues using specificity-determining residue (SDR)-grafting and de-immunization methods. The resultant humanized antibody, HzS-III, had the same affinity and epitope specificity as HuS10 and had reduced immunogenic potential, as assessed by T-cell epitope analysis. Thus, SDR grafting in combination with de-immunization may be a useful strategy for minimizing the immunogenicity of humanized antibodies. In addition, HzS-III may be a good candidate for immunoprophylaxis of HBV infection.

Gene and microRNA expression signatures of human mesenchymal stromal cells in comparison to fibroblasts.

Human mesenchymal stromal cells (MSCs) offer great hope for the treatment of tissue degenerative and immune diseases, but their phenotypic similarity to dermal fibroblasts may hinder robust cell identification and isolation from diverse tissue harvests. To identify genetic elements that can reliably discriminate MSCs from fibroblasts, we performed comparative gene and microRNA expression profiling analyses with genome-wide oligonucleotide microarrays. When taken globally, both gene and microRNA expression profiles of MSCs were highly similar to those of fibroblasts, accounting well for their extensive phenotypic and functional overlaps. Scattered expression differences were pooled to yield an MSC-specific molecular signature, consisting of 64 genes and 21 microRNAs whose expressions were at least 10-fold and two-fold higher, respectively, in MSCs compared with fibroblasts. Genes either encoding transmembrane proteins or associated with tumors were relatively abundant in this signature. These data should provide the molecular basis not only for the discovery of novel diagnostic markers discriminating MSCs from fibroblasts, but also for further studies on MSC-specific signaling mechanisms.

Cytokine secretion profiling of human mesenchymal stem cells by antibody array.

Mesenchymal stem cells (MSCs) provide not only cell sources for connective tissues but also the control of hematopoiesis and immune response. A multitude of cytokines and growth factors secreted from MSCs are known to confer such multifunctional functionality, but their overall spectrum and the respective expression strength have not been thoroughly illustrated. In this study, we have obtained the comprehensive cytokine secretion profile of human bone marrow (BM)-derived MSCs, with the use of an antibody array recognizing 120 cytokines and chemokines. The array membrane incubated with the secretion media of the cells featured a predominant hybridization signal for IL-6 and moderately elevated signals for IL-8, TIMP-2, MCP-1, VEGF and OPG. This cytokine secretion profile was found to be common to all cell lines from three different donors, and also similar but not identical to that of umbilical cord blood-derived cells, suggesting that the trophic nature of the MSCs might depend slightly on the cell origin but not on individuality of the donors. Our results here may provide the molecular basis for further studies on MSC-assisted biological processes, such as connective tissue homeostasis, hematopoiesis and immune modulation.

Selection of an affinity-matured antibody against a defined epitope by phage display of an immune antibody library.

In a previous study, we generated a murine hepatitis B virus (HBV)-neutralizing monoclonal antibody (mAb), KR127, that binds to an epitope (amino acids 37-45, NSNNPDWDF) of the preS1 antigen. Furthermore, an epitope tag, S1 (NANNPDWDF), was developed for protein tagging. The aim of the present study was to develop a high-affinity antibody to the same preS1 epitope. Mice were immunized with the N-terminal domain of human thrombopoietin fused to the S1 tag (nTPO-S1), and a phage-displayed chimeric Fab library was constructed and screened by panning against nTPO-S1. A high-affinity antibody (3-34) was selected that binds to the preS1 antigen. The IgG molecules of 3-34 showed approximately nine-fold higher affinity (K(D) 1.2 nM) for preS1 compared with KR127 (K(D) 10.4 nM), competed with KR127 for binding to the epitope, and bound to HBV particles. This study provides a simple and efficient way to develop a high-affinity antibody to a defined epitope by phage display of an immune antibody library.

Monoclonal anti-thrombopoietin antibodies generated by genetic immunization.

Thrombopoietin (TPO) is a megakaryocyte growth and differentiation factor that is currently being investigated as a therapeutic for cancer patients undergoing myelosuppressive chemotherapy. We generated monoclonal antibodies (MAbs) specific for human thrombopoietin (hTPO) by genetic immunization using an hTPO expression plasmid and an adjuvant plasmid that encodes mouse granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). All genetically immunized mice exhibited a high humoral immune response. Splenocytes from these mice were used to generate hybridomas. Two MAbs, designated 2B9A10 and 4C16B15 (of IgG1 and IgG3 isotypes, respectively), were subsequently selected and produced. They specifically recognized and precipitated recombinant hTPO produced by mammalian cells and were effective in sandwich enzyme-linked immunosorbent assays (ELISAs) for hTPO quantitation. Our results demonstrate that these MAbs should be useful for purification and quantitation of hTPO in clinical and laboratory settings.

Construction, affinity maturation, and biological characterization of an anti-tumor-associated glycoprotein-72 humanized antibody.

The tumor-associated glycoprotein (TAG)-72 is expressed in the majority of human adenocarcinomas but is rarely expressed in most normal tissues, which makes it a potential target for the diagnosis and therapy of a variety of human cancers. Here we describe the construction, affinity maturation, and biological characterization of an anti-TAG-72 humanized antibody with minimum potential immunogenicity. The humanized antibody was constructed by grafting only the specificity-determining residues (SDRs) within the complementarity-determining regions (CDRs) onto homologous human immunoglobulin germ line segments while retaining two mouse heavy chain framework residues that support the conformation of the CDRs. The resulting humanized antibody (AKA) showed only about 2-fold lower affinity compared with the original murine monoclonal antibody CC49 and 27-fold lower reactivity to patient serum compared with the humanized antibody HuCC49 that was constructed by CDR grafting. The affinity of AKA was improved by random mutagenesis of the heavy chain CDR3 (HCDR3). The highest affinity variant (3E8) showed 22-fold higher affinity compared with AKA and retained the original epitope specificity. Mutational analysis of the HCDR3 residues revealed that the replacement of Asn(97) by isoleucine or valine was critical for the affinity maturation. The 3E8 labeled with (125)I or (131)I showed efficient tumor targeting or therapeutic effects, respectively, in athymic mice with human colon carcinoma xenografts, suggesting that 3E8 may be beneficial for the diagnosis and therapy of tumors expressing TAG-72.

An anthrax lethal factor-neutralizing monoclonal antibody protects rats before and after challenge with anthrax toxin.

Lethal factor (LF) is a component of anthrax lethal toxin (LeTx). We generated anti-LF murine monoclonal antibodies (MAbs) that show LeTx-neutralizing activity in vitro and in vivo. Anti-LF MAbs were generated by immunization with recombinant LF, and the MAbs showing LeTx-neutralizing activity in vitro were selected. Two MAbs with the highest affinities, 5B13B1 (dissociation constant [K(d)], 2.62 nM) and 3C16C3 (K(d), 8.18 nM), were shown to recognize the same or closely overlapping epitopes on domain III of LF. The 50% inhibitory concentration of 5B13B1 (0.21 microg/ml) was approximately one-third that of 3C16C3 (0.63 microg/ml) in the in vitro LeTx-neutralization assay. The 5B13B1 antibody, which had the highest neutralizing activity, provided perfect protection against LeTx challenge in an in vivo LeTx neutralization assay using Fisher 344 rats. In addition, the antibody showed pre- and postexposure prophylactic effects in the animal experiments. This is the first report that an MAb binding to domain III of LF has neutralizing activity against LeTx. The 5B13B1 antibody may be useful in prophylaxis against anthrax poisoning.

Neutralizing human monoclonal antibodies to hepatitis A virus recovered by phage display.

Four human monoclonal antibodies (MAbs) to hepatitis A virus (HAV) were isolated from a phage-displayed antibody library constructed from the peripheral blood lymphocytes (PBLs) of HAV-immune donors. The four MAbs showed differences in their affinity: two (HA6, HA9) of them were dominant after four rounds of panning, and showed higher affinity than the other two (HA1, HA12). All four MAbs showed HAV-neutralizing activity in radioimmunofocus inhibition assay and their neutralizing activity was positively correlated with their affinities. Analysis of their epitope specificity by cross-competition binding assays suggested that HA6 and HA9 recognize extensively overlapping epitopes, which overlap with those of HA1 and HA12, although HA1 and HA12 recognize distinct epitopes. In addition, competition assays with known neutralizing murine MAbs suggested that the epitopes of four human MAbs extensively overlap with those of B5B3 and K34C8 which are distinct but reside within the single, immunodominant neutralization site on the HAV capsid. The human MAbs (HA6 and HA9) with highest affinity may be useful in the immunoprophylaxis of HAV infection.