A mass and charge balanced metabolic model of Setaria viridis revealed mechanisms of proton balancing in C4 plants.

BACKGROUND: C4 photosynthesis is a key domain of plant research with outcomes ranging from crop quality improvement, biofuel production and efficient use of water and nutrients. A metabolic network model of C4 "lab organism" Setaria viridis with extensive gene-reaction associations can accelerate target identification for desired metabolic manipulations and thereafter in vivo validation. Moreover, metabolic reconstructions have also been shown to be a significant tool to investigate fundamental metabolic traits. RESULTS: A mass and charge balance genome-scale metabolic model of Setaria viridis was constructed, which was tested to be able to produce all major biomass components in phototrophic and heterotrophic conditions. Our model predicted an important role of the utilization of NH[Formula: see text] and NO[Formula: see text] ratio in balancing charges in plants. A multi-tissue extension of the model representing C4 photosynthesis was able to utilize NADP-ME subtype of C4 carbon fixation for the production of lignocellulosic biomass in stem, providing a tool for identifying gene associations for cellulose, hemi-cellulose and lignin biosynthesis that could be potential target for improved lignocellulosic biomass production. Besides metabolic engineering, our modeling results uncovered a previously unrecognized role of the 3-PGA/triosephosphate shuttle in proton balancing. CONCLUSIONS: A mass and charge balance model of Setaria viridis, a model C4 plant, provides the possibility of system-level investigation to identify metabolic characteristics based on stoichiometric constraints. This study demonstrated the use of metabolic modeling in identifying genes associated with the synthesis of particular biomass components, and elucidating new role of previously known metabolic processes.

Analgesic and sedative effects of intranasal dexmedetomidine in third molar surgery under local anaesthesia.

BACKGROUND: Dexmedetomidine (DEX) is an alpha 2-adrenoreceptor agonist, which induces sedation and analgesia. This study aimed to determine whether intranasal DEX offered perioperative sedation and better postoperative analgesia. METHODS: Patients having unilateral third molar surgery under local anaesthesia were recruited and allocated to receive either intranasal DEX 1 µg kg(-1) (Group D) or same volume of saline (Group P) 45 min before surgery. Patient-controlled sedation with propofol was offered as a rescue sedative. Perioperative sedation, postoperative pain relief and analgesic consumption, vital signs, adverse events, postoperative recovery, and satisfaction in sedation and analgesia were assessed. RESULTS: Thirty patients from each group were studied. Areas under curve (AUC) of postoperative numerical rating scale (NRS) pain scores 1-12 h at rest and during mouth opening were significantly lower in Group D (P=0.003 and 0.009, respectively). AUC BIS values and OAA/S sedation scores were significantly lower before surgery and at the recovery area (all P<0.01) with significantly less intra-operative propofol used in group D (P<0.01). In group D, heart rate was significantly lower at recovery period (P=0.005) while systolic blood pressure in different periods of the study (all P<0.01), but the decreases did not require treatment. More patients from placebo group experienced dizziness (P=0.026) but no serious adverse event was found. No difference was found in postoperative psychomotor recovery and satisfaction in pain relief and sedation. CONCLUSIONS: Patients receiving intranasal DEX for unilateral third molar surgery with local anaesthesia were more sedated perioperatively with better postoperative pain relief. No delay in psychomotor recovery was seen.

A study to assess the value of bispectral analysis in intravenous sedation with midazolam during third molar surgery under local anaesthesia.

This study aimed to determine whether bispectral index (BIS) can be used as an indicator of sedation and recovery with intravenous midazolam. In Part A, 30 healthy patients undergoing third molar extraction under local anaesthesia were recruited. They were sedated with intravenous midazolam titrated to clinical endpoints. BIS values were recorded when adequately sedated (BIS(S)) and when clinical recovery criteria were met (BIS). In Part B, another 30 patients were sedated to the range of BIS(S) obtained in Part A. Recovery was assessed postoperatively when the range of BIS(R) from Part A was reached. BIS titrated patients required less midazolam (p < 0.001). Seventy percent of Part B patients required increments of midazolam during surgery, compared to 16.7% in Part A (p < 0.001). Total dose of midazolam given was lower in Part B (p = 0.025). BIS is not effective as a sole indicator of endpoint in sedation with intravenous midazolam.

Developing a short form of Oral Health Impact Profile (OHIP) for dental aesthetics: OHIP-aesthetic.

OBJECTIVES: To develop and evaluate shortened forms of the Oral Health Impact Profile (OHIP) for discriminating dental aesthetics problems and evaluating dental aesthetic outcomes. METHODS: Eighty-seven subjects self-completed the 49-item OHIP at baseline and 63 at follow up (8 weeks later), with the intervention of applying an array of tooth-whitening products. Expert-based approach and regression analysis (on baseline data) were undertaken to derive two subset questionnaires (OHIP-conceptual and OHIP-regression). Their discriminatory ability for dental aesthetics and their responsiveness to tooth whitening were compared with the original OHIP-49, Slade's OHIP-14 and a Chinese short-form version of OHIP. RESULTS: The measures developed were strongly associated with self-rating of dental aesthetics (P < 0.001) unlike OHIP-49 (P = 0.03) or other OHIP short forms (P > 0.05). The measures were also reliable (Cronbach's alpha 0.86) and comparable with the other OHIP forms. In terms of effect size, OHIP-conceptual was more effective in measuring changes than the one based on the regression analysis, the original OHIP-49, OHIP-14 and the Chinese version of the short-form OHIP. It also exhibited a less susceptibility to floor effects than other OHIP forms. CONCLUSION: A modified short form of the OHIP derived (OHIP-conceptual) was the most favorable in discriminating dental aesthetics, was reliable and most sensitive to the dental aesthetics intervention - tooth whitening.

The sensitivity and responsiveness of an oral health related quality of life measure to tooth whitening.

OBJECTIVES: To assess the sensitivity and responsiveness of an oral health related quality of life measure to tooth whitening. METHODS: Following screening at a clinic, 87 subjects were given an array of tooth whitening products to use at home and reviewed 8 weeks later. Subjects self-completed the 49-item Oral Health Impact Profile (OHIP) at baseline and follow-up, and rated their satisfaction with the whiteness of their teeth compared to baseline on a global transition scale. RESULTS: In terms of sensitivity, observed changes were apparent in overall OHIP scores (P<0.05) and across several domains, notably functional limitation (P<0.01). However, the magnitude of change (effect size) was generally small except for the functional domain. There was an observed gradient in observed change in OHIP scores and in the magnitude of such changes (effect sizes) in relation to global rating of satisfaction with the outcome, supporting the responsiveness of the measure. CONCLUSION: The OHIP scale is sensitive and responsive to the effects of tooth whitening. Greatest sensitivity and responsiveness was in relation to functional limitations. These findings have implications for the use of oral health related quality of life measures as an outcome measure of interventions aimed at improving dental aesthetics through tooth whitening.

Wear performance of ultrahigh molecular weight polyethylene/quartz composites.

Ultrahigh molecular weight polyethylene (UHMWPE)/quartz composites were compression molded in the presence of organosiloxane, and then hydrolyzed. The used organosiloxane is vinyl tri-ethyloxyl silane. The gelation, the melting behavior, the crystallinity, the mechanical properties and the wear resistance of UHMWPE/quartz composites were investigated. The results showed that organosiloxane can act as a cross-linking agent for UHMWPE matrix and serve as a coupling agent for improving the bonding between the quartz particles and the UHMWPE matrix. The correlation between the various properties and the morphology of the composites has been discussed. At about 0.5phr organsiloxane while the degree of crystallinity of the composite is at the peak value of 57%, the mechanical properties and the wear resistance of UHMWPE/quartz composites reaches their maximum.

Molecular engineering of proteins and polymers for targeting and intracellular delivery of therapeutics.

There are many protein and DNA based therapeutics under development in the biotechnology and pharmaceutical industries. Key delivery challenges remain before many of these biomolecular therapeutics reach the clinic. Two important barriers are the effective targeting of drugs to specific tissues and cells and the subsequent intracellular delivery to appropriate cellular compartments. In this review, we summarize protein engineering work aimed at improving the stability and refolding efficiency of antibody fragments used in targeting, and at constructing new streptavidin variants which may offer improved performance in pre-targeting delivery strategies. In addition, we review recent work with pH-responsive polymers that mimic the membrane disruptive properties of viruses and toxins. These polymers could serve as alternatives to fusogenic peptides in gene therapy formulations and to enhance the intracellular delivery of protein therapeutics that function in the cytoplasm.

What is the relationship between the Glasgow coma scale and airway protective reflexes in the Chinese population?

AIM: To describe the relationship of gag and cough reflexes to Glasgow coma score (GCS) in Chinese adults requiring critical care. METHOD: Prospective observational study of adult patients requiring treatment in the trauma or resuscitation rooms of the Emergency Department, Prince of Wales Hospital, Hong Kong. A long cotton bud to stimulate the posterior pharyngeal wall (gag reflex) and a soft tracheal suction catheter were introduced through the mouth to stimulate the laryngopharynx and elicit the cough reflex. Reflexes were classified as normal, attenuated or absent. RESULTS: A total of 208 patients were recruited. Reduced gag and cough reflexes were found to be significantly related to reduced GCS (p=0.014 and 0.002, respectively). Of 33 patients with a GCS≤8, 12 (36.4%) had normal gag reflexes and 8 (24.2%) had normal cough reflexes. 23/62 (37.1%) patients with a GCS of 9-14 had absent gag reflexes, and 27 (43.5%) had absent cough reflexes. In patients with a normal GCS, 22.1% (25/113) had absent gag reflexes and 25.7% (29) had absent cough reflexes. CONCLUSIONS: Our study has shown that in a Chinese population with a wide range of critical illness (but little trauma or intoxication), reduced GCS is significantly related to gag and cough reflexes. However, a considerable proportion of patients with a GCS≤8 have intact airway reflexes and may be capable of maintaining their own airway, whilst many patients with a GCS>8 have impaired airway reflexes and may be at risk of aspiration. This has important implications for airway management decisions.

A comparison of dexmedetomidine and midazolam for sedation in third molar surgery.

This randomised, double-blind study compared dexmedetomidine and midazolam for intravenous sedation during third molar surgery under local anaesthesia. Sixty patients received either dexmedetomidine (up to 1 microg x kg(-1)) or midazolam (up to 5 mg), which was infused until the Ramsay Sedation Score was four or the maximum dose limit was reached. Intra-operative vital signs, postoperative pain scores and analgesic consumption, amnesia, and satisfaction scores for patients and surgeons, were recorded. Sedation was achieved by median (IQR (range)) doses of 47 microg (39-52 (25-76)) or 0.88 microg x kg(-1) (0.75-1.0 (0.6-1.0)) dexmedetomidine, and 3.6 mg (3.3-4.4 (1.9-5.0)) or 0.07 mg x kg(-1) (0.055-0.085 (0.017-0.12)) midazolam. Heart rate and blood pressure during surgery were lower in dexmedetomidine group. There was no significant difference in satisfaction or pain scores. Midazolam was associated with greater amnesia. Dexmedetomidine produces comparable sedation to midazolam.

'Smart' delivery systems for biomolecular therapeutics.

OBJECTIVE: There is a strong need for drug delivery systems that can deliver biological signals from biomaterials and tissue engineering scaffolds, and a particular need for new delivery systems that can efficiently deliver biomolecules to intracellular targets. Viruses and pathogens have evolved potent molecular machinery that sense the lowered pH gradient of the endosomal compartment and become activated to destabilize the endosomal membrane, thereby enhancing protein or DNA transport to the cytoplasmic compartment. A key feature of many of these biological delivery systems is that they are reversible, so that the delivery systems are not directly toxic. These delivery systems have the ability to change their structural and functional properties and thus display remarkable 'smart' material properties. The objective of this presentation is to review the initial development of smart polymeric carriers that mimic these biological delivery systems and combine similar pH-sensitive, membrane-destabilizing activity for the delivery of therapeutic biomolecules. DESIGN: We have developed new 'smart' polymeric carriers to more effectively deliver and broaden the available types of biomolecular therapeutics. The polymers are hydrophilic and stealth-like at physiological pH, but become membrane-destabilizing after uptake into the endosomal compartment where they enhance the release of therapeutic cargo into the cytoplasm. They can be designed to provide a range of pH profiles and membrane-destabilizing activities, allowing their molecular properties to be matched to specific drugs and loading ranges. A versatile set of linker chemistries is available to provide degradable conjugation sites for proteins, nucleic acids, and/or targeting moieties. RESULTS: The physical properties of several pH-responsive polymers were examined. The activity and pH profile can be manipulated by controlling the length of hydrophobic alkyl segments. The delivery of poly(propyl acrylic acid) (PPAA)-containing lipoplexes significantly enhanced wound healing through the interconnected effects of altered extracellular matrix organization and greater vascularization. PPAA has also been shown to enhance cytoplasmic delivery of a model protein therapeutic. Polymeric carriers displaying pH-sensitive, membrane-destabilizing activity were also examined. The pH profile is controlled by the choice of the alkylacrylic acid monomer and by the ratio of the carboxylate-containing alkylacrylic acid monomer to alkylacrylate monomer. The membrane destabilizing activity is controlled by the lengths of the alkyl segment on the alkylacrylic acid monomer and the alkylacrylate monomer, as well as by their ratio in the final polymer chains. CONCLUSION: The molecular mechanisms that proteins use to sense and destabilize provide interesting paradigms for the development of new polymeric delivery systems that mimic biological strategies for promoting the intracellular delivery of biomolecular drugs. The key feature of these polymers is their ability to directly enhance the intracellular delivery of proteins and DNA, by destabilizing biological membranes in response to vesicular compartment pH changes. The ability to deliver a wide variety of protein and nucleic acid drugs to intracellular compartments from tissue engineering and regenerative scaffolds could greatly enhance control of important processes such as inflammation, angiogenesis, and biomineralization.

A pH-sensitive polymer that enhances cationic lipid-mediated gene transfer.

The efficient release of nonviral gene carriers from endosomes is an important step for the successful delivery of DNA into the cell nucleus. A synthetic pH-sensitive anionic polymer, poly(propylacrylic acid) (PPAA), was designed to aid in endosomal escape of nonviral vectors and improve the transfection efficiencies with these vectors. Transfection of NIH3T3 fibroblasts with ternary physical mixtures of the cationic lipid DOTAP, pCMVbeta plasmid DNA, and PPAA showed marked enhancement of both gene expression levels and fraction of cells transfected compared to binary control mixtures of DOTAP and DNA. PPAA also significantly improved the serum-stability of DOTAP/DNA vectors. The DOTAP/DNA/PPAA vectors maintained high levels of transfection in media containing up to 50% serum. The striking enhancement of transfection efficiency with cationic lipid/DNA/PPAA mixtures, along with the enhanced serum-stability, suggests that PPAA may provide significant improvements for the in vivo intracellular delivery of drugs such as DNA, oligonucleotides, proteins, and peptides.

Impaired G(s)alpha and adenylyl cyclase cause beta-adrenoceptor desensitization in chronically hypoxic rat hearts.

The effects of beta-adrenoceptor stimulation with isoproterenol on electrically induced contraction and intracellular calcium ([Ca(2+)](i)) transient, and cAMP in myocytes from both hypertrophied right and nonhypertrophied left ventricles of rats exposed to 10% oxygen for 4 wk, were significantly attenuated. The increased [Ca(2+)](i) transient in response to cholera toxin was abolished, whereas increased cAMP after NaF significantly attenuated. The biologically active isoform, G(s)alpha-small (45 kDa), was reduced while the biologically inactive isoform, G(s)alpha-large (52 kDa), increased. The increased electrically induced [Ca(2+)](i) transient and cAMP with 10-100 microM forskolin were significantly attenuated in chronically hypoxic rats. The content of G(i)alpha(2), the predominant isoform of G(i) protein in the heart, was unchanged. Results indicate that impaired functions of G(s) protein and adenylyl cyclase cause beta-adrenoceptor desensitization. The impaired function of the G(s) protein may be due to reduced G(s)alpha-small and/or increased G(s)alpha-large, which does not result from changes in G(i) protein. Responses to all treatments were the same for right and left ventricles, indicating that the impaired cardiac functions are not secondary to cardiac hypertrophy.

Founder's Award, Society for Biomaterials. Sixth World Biomaterials Congress 2000, Kamuela, HI,May 15-20, 2000. Really smart bioconjugates of smart polymers and receptor proteins.

Over the past 18 years we have been deeply involved with the synthesis and applications of stimuli-responsive polymer systems, especially polymer-biomolecule conjugates. This article summarizes our work with one of these conjugate systems, specifically polymer-protein conjugates. We include conjugates prepared by random polymer conjugation to lysine amino groups, and also those prepared by site-specific conjugation of the polymer to specific amino acid sites that are genetically engineered into the known amino acid sequence of the protein. We describe the preparation and properties of thermally sensitive random conjugates to enzymes and several affinity recognition proteins. We have also prepared site-specific conjugates to streptavidin with temperature-sensitive polymers, pH-sensitive polymers, and light-sensitive polymers. The preparation of these conjugates and their many fascinating applications are reviewed in this article.