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We investigate offline dynamic optimization (ODO) and multi-layer control schemes to optimally operate the epoxidation of oleic acid. The process involves a two-phase reaction mixture with highly exothermic reactions wherein the reactor temperature needs to be controlled to guarantee safe operation. Optimal operating conditions are computed using ODO and the control scheme is responsible to enforce such conditions. We compare a conventional control scheme, which uses PI controllers, against two multi-layer control schemes: (i) using nonlinear model predictive control (NMPC) and (ii) using dynamic real-time optimization (DRTO) together with NMPC. With ODO and NMPC we achieve a reduction in process time greater than 75%, compared to a standard process, while still maintaining safe operating conditions. With DRTO and NMPC, the same reduction in process time is obtained, however with a maximum violation of the temperature constraint by 12%. This may be due to model-plant mismatch and large computational time required for the optimization.

Background Combination of immune checkpoint inhibitors and mitogen-activated protein kinase (MAPK) pathway inhibitors (MAPKi) has been proposed to enhance the durability of anti-tumour responses induced by MAPKi. Here, we present phase I safety results from an open-label, phase I/II study of pembrolizumab (PEM), encorafenib (ENC) and binimetinib (BIN) triplet therapy in advanced, B-Raf proto-oncogene serine/threonine kinase (BRAF)V600–mutated melanoma (IMMU-TARGET, NCT02902042 ). Methods The dose finding phase I part used a 3 + 3 design, starting with the approved doses of PEM (200 mg every three weeks), ENC (450 mg once daily [QD]) and BIN (45 mg twice daily [BID]) as dose level (DL) 0. Reduction of the ENC and BIN doses (300 mg QD and 30 mg BID at DL-1 and 200 mg QD and 30 mg BID at DL-2) was preplanned in case of ≥2 dose-limiting toxicities (DLTs). Primary objectives were to estimate the recommended phase II dose of the triplet combination, DLT and safety. As per the sponsor's decision, the study was terminated after the phase I part, as the clinical efficacy of the combination is currently being investigated in a pivotal, placebo-controlled (PEM mono), double-blinded phase III trial (STARBOARD, NCT04657991 ). Results Fifteen patients were enrolled. DLTs of DL0 were creatine phosphokinase (CPK) elevation plus cytokine release syndrome (n = 1) and gamma glutamyl transferase (GGT) increase (n = 1). No DLT was observed in further 3 + 3 patients at DL-1. One (isolated GGT elevations) DLT of DL0 was questionable, as the patient had further episodes of isolated GGT elevations after treatment discontinuation. Hence, further 6 patients were enrolled at DL0: here, no DLT occurred. In total, 13 of 15 patients (87%) experienced a treatment-related adverse event (TRAE) and 8 patients (53%), a grade ≥III TRAE; there were no TRAE-related deaths. Increases in aspartate aminotransferases, GGT (6/15 patients) and CPK elevations (4/15) were the most common grade III–IV TRAE. In median, patients received triplet therapy for 24 weeks (interquartile range [IQR], 12–45). Of the 14 patients evaluable for efficacy, the overall response rate was 64% (95% confidence interval [CI], 35–87). At a median follow-up of 25 months (IQR, 9–28), progression-free survival at 12 months was 41% (95% CI, 13–68). Conclusions Triplet therapy with PEM, ENC and BIN as used in the study was feasible and safe and led to clinically meaningful disease control.

Objective Increasing the amounts of functionally competent brown adipose tissue (BAT) in adult humans has the potential to restore dysfunctional metabolism and counteract obesity. In this study, we aimed to characterize the human perirenal fat depot, and we hypothesized that there would be regional, within-depot differences in the adipose signature depending on local sympathetic activity. Methods We characterized fat specimens from four different perirenal regions of adult kidney donors, through a combination of qPCR mapping, immunohistochemical staining, RNA-sequencing, and pre-adipocyte isolation. Candidate gene signatures, separated by adipocyte morphology, were recapitulated in a murine model of unilocular brown fat induced by thermoneutrality and high fat diet. Results We identified widespread amounts of dormant brown adipose tissue throughout the perirenal depot, which was contrasted by multilocular BAT, primarily found near the adrenal gland. Dormant BAT was characterized by a unilocular morphology and a distinct gene expression profile, which partly overlapped with that of subcutaneous white adipose tissue (WAT). Brown fat precursor cells, which differentiated into functional brown adipocytes were present in the entire perirenal fat depot, regardless of state. We identified SPARC as a candidate adipokine contributing to a dormant BAT state, and CLSTN3 as a novel marker for multilocular BAT. Conclusions We propose that perirenal adipose tissue in adult humans consists mainly of dormant BAT and provide a data set for future research on factors which can reactivate dormant BAT into active BAT, a potential strategy for combatting obesity and metabolic disease., Graphical abstract Image 1, Highlights • Dormant brown adipose tissue (BAT) dominates the perirenal adipose depot of adult humans. • Multilocular BAT accumulates adjacent to local sympathetic sources. • Dormant BAT displays a transcriptomic signature distinct from multilocular BAT and white adipose tissue. • Brown fat precursor cells are present in human dormant perirenal BAT. • SPARC is associated with a dormant BAT phenotype.

Background This study evaluated whether a multi-ingredient surfactant colloidal silver technology was noninferior to a 4% chlorhexidine gluconate (CHG) antiseptic on immediate and persistent antimicrobial activity. Methods The inguinal regions of 81 healthy adults were demarcated into 4 quadrants, and 3 were used for testing each product at baseline, 10 minutes, and 6 hours postapplication. The log of the number of colony forming units was obtained using a cylinder sampling technique. The 95% confidence interval of the test product to the control product with a margin of 0.65 was established as the upper limit of noninferiority. Results A total of 81 individuals were enrolled. The colloidal silver product was found to be noninferior to 4% CHG at both 10 minutes and 6 hours postapplication. Conclusions The colloidal silver-based product was noninferior to the 4% CHG product at 10 minutes and 6 hours postapplication.

Sleep benefits memory in young adults, and this effect may be particularly strong for representations associated with negative emotion. Many aspects of sleep important for memory consolidation change with aging, particularly by middle age, suggesting that sleep-related consolidation may be reduced. However, the influence of sleep on memory has rarely been investigated in a middle-aged population. In the current study, young and middle-aged adults viewed negative and neutral pictures and underwent a recognition test after sleep or wake. Subjective emotional reactivity was also measured. Compared to waking, sleep benefited memory in young adults. Performance did not differ between sleep and wake groups in middle-aged adults, and it matched the level of young adults who slept. The effect of sleep versus wake was not influenced by memory valence in either age group. These results suggest the relative influence of sleep compared to wake on memory declines with aging, specifically by middle age, and that this decline extends to negative memory.

Rationale Little is known about how perinatal and childhood factors influence lung function change between childhood and adolescence. Objectives To investigate possible early life predictors of change in FEV1 between age 8 and 16 years. In addition, to investigate possible predictors of having persistently low lung function (FEV1 Methods The BAMSE birth cohort study collected data throughout childhood on environmental factors, individual characteristics, and spirometric measures at 8 and 16 years (n = 1425). Associations between early life predictors (n = 31) and FEV1 increase between 8 and 16 years were assessed with linear regression. Predictors of having persistently low lung function were examined. Results Few factors were consistently associated with altered lung function growth, although low birth weight, asthma heredity (paternal), secondhand smoke in infancy, and season of birth had a significant impact (p-value ≤0.01). The majority of subjects stayed however within the same category of lung function between ages 8 and 16 years (in total 821/1425 = 58%). Predictors associated with having persistently low lung function were gestational age, secondhand smoke (at 2 and 8 years of age), and factors related to lower respiratory tract infections in infancy. Conclusions In summary, rather few exposures in childhood were identified to have a significant impact on lung function growth between childhood and adolescence. Our data support previous study findings indicating that lung function development is influenced by factors before birth and in infancy, including second hand tobacco smoke.

Low oxygen concentration or hypoxia plays a crucial role in the diagnosis and treatment of tumours and cancer. Although there are some existing techniques, such as proton magnetic resonance imaging (MRI), for the detection of molecular oxygen in tissues, new non-invasive techniques for directly measuring the extent and spatial distribution of hypoxia in cancerous or tumorous tissues are still needed. In this study, we propose a new non-invasive method using muons that are available in accelerator facilities, such as the Japan Proton Accelerator Research Complex in Japan and the Rutherford Appleton Laboratory in the UK. Unlike existing MRI methods, the proposed muon-based method can be applied without any external magnetic field. With the help of muonium response in solutions, we measured oxygen concentration in the aqueous solutions of various biological materials, such as haemoglobin, albumin, serum, and tris-buffered saline, under controlled molecular oxygen concentrations. The observed relaxation rate pattern of muonium through a fixed oxygen concentration sample and that with a sample concentration of less than 1% oxygen shows that this method may be applicable in detecting molecular oxygen levels in various tissues of the human body. Based on our experimental results, we have estimated the expected muonium spectra in normal physiological concentrations of haemoglobin in blood. Further systematic studies on other biological solutions are required before a potential clinical application of the method is devised.

There has been some evidence that a short heating exposure of the testicles can lead to temporary sterility. However, little attention has been paid to analyzing the temperature/time pattern throughout the testicle and how metabolism and blood perfusion affect the temperature profile. We use the Penne bioheat equation that accounts for the elements of conduction, perfusion, and metabolism along with the COMSOL Finite Element Method for the analysis of this thermoregulation problem. Typically, the testicles were modeled as two uniform spheres and exposed to an increase in surface temperature of 42 °C for 30 min. We found that heat produced by tissue metabolism had a minor effect on the temperature pattern. However, blood flow from the core (at 37 °C) could prevent the region where the testicles touch each other from reaching the target temperature of 42 °C. Further, we found that for animals, as rodents and monkeys that are used for contraception studies, the temperature distribution on heating testicles is much more uniform than for humans. Thus, extrapolation results from animal models to humans should be done with caution.

Objective MicroRNAs (miRNAs) are increasingly recognized as fine-tuning regulators of metabolism, and are dysregulated in several disease conditions. With their capacity to rapidly change gene expression, miRNAs are also important regulators of development and cell differentiation. In the current study, we describe an impaired myogenic capacity of muscle stem cells isolated from humans with type 2 diabetes (T2DM) and assess whether this phenotype is regulated by miRNAs. Methods We measured global miRNA expression during in vitro differentiation of muscle stem cells derived from T2DM patients and healthy controls. Results The mir-23b/27b cluster was downregulated in the cells of the patients, and a pro-myogenic effect of these miRNAs was mediated through the p53 pathway, which was concordantly dysregulated in the muscle cells derived from humans with T2DM. Conclusions Our results indicate that we have identified a novel pathway for coordination of myogenesis, the miR-23b/27b-p53 axis that, when dysregulated, potentially contributes to a sustained muscular dysfunction in T2DM., Graphical abstract Image 1, Highlights • miR-23b and miR-27b are pro-myogenic and are downregulated in T2DM. • miR-23b and miR-27b regulate myogenesis through the p53 pathway. • The p53 pathway is concordantly dysregulated in T2DM.

Bacterial biofilms are surface-attached communities of slow-growing or non-replicating bacteria tolerant to conventional antibiotic therapies. Although biofilms are known to occur in ca. 80% of all bacterial infections, no therapeutic agent has been developed to eradicate bacteria housed within biofilms. We have discovered that nitroxoline, an antibacterial agent used to treat urinary tract infections, displays broad-spectrum biofilm-eradicating activities against major human pathogens, including drug-resistant Staphylococcus aureus and Acinetobacter baumannii strains. In this study, the effectiveness of nitroxoline to eradicate biofilms was determined using an in vitro [minimum biofilm eradication concentration (MBEC) = 46.9 µM against A. baumannii] and ex vivo porcine skin model (2-3 log reduction in viable biofilm cells). Nitroxoline was also found to eradicate methicillin-resistant S. aureus (MRSA) persister cells in non-biofilm (stationary) cultures, whereas vancomycin and daptomycin were found to be inactive. These findings could lead to effective, nitroxoline-based therapies for biofilm-associated infections.

Highlights • School employee health and well-being directly impact student learning and success. • Higher physical activity levels were associated with better cardiometabolic health. • High stress had a negative impact on the relationship between PA and BMI. • Stress may affect heart health, potentially through an impact on health behaviors. • Enhancing school employee well-being may improve student academic achievement., Among school employees, it has been reported that poor physical and mental health, as well as high stress and large workloads, have resulted in high absenteeism and low retention. The consequences of unhealthy behaviors and stress can extend to students, impacting academic achievement and school costs. Our objective was to examine the impact of school employees’ physical activity (PA), diet quality and perceived occupational stress on cardiometabolic health, and explore how stress may influence the impact of PA and diet on health. In this cross-sectional study, employees from lower-income Massachusetts schools participated in Wellness Assessments (2015–2016), including measured height, weight, and lipids [total (TC), high-density lipoprotein (HDL-C), low-density lipoprotein (LDL-C)]. Self-administered surveys were used to collect demographic, stress, PA and 24-hour food intake data. Linear regression models were used to examine the relationship among health behaviors (PA and diet), stress and cardiometabolic health. An interaction between stress and health behaviors was also explored. Seventy-four employees (66% teachers) participated. Overweight/obesity (mean BMI: 25.6 kg/m2), high TC and LDL-C were observed in 47%, 4%, and 34%, respectively, and moderate-to-vigorous PA (MVPA) was low (median: 17 min/day). Positive associations were identified between MVPA and cardiometabolic health, but not diet. The effect of MVPA on BMI was modified by stress (p-for-interaction = 0.001), with higher levels of stress associated with a diminished protective association between MVPA and BMI. Higher levels of PA were associated with more favorable cardiometabolic health, with increasing levels of stress minimizing the beneficial effect of PA on BMI.

Introduction Afatinib is a tyrosine kinase inhibitor (TKI), that has been approved for treating patients with epidermal growth factor receptor (EGFR) mutated advanced non-small-cell lung cancer (NSCLC). Stevens-Johnson syndrome (SJS) related to EGFR directed TKIs is a rare adverse event. Case presentation We report a case of a 79-year-old white female with EGFR-mutated, metastatic non-small-cell lung cancer treated with afatinib as first-line palliative treatment, who developed a SJS after two months of treatment. Discontinuation of the TKI and systemic glucocorticoid treatment led to improvement of symptoms and recovery. Conclusion Severe adverse cutaneous drug reactions that predominantly involve the skin and mucous membranes during treatment with afatinib should alert clinicians to suspect SJS and react appropriately.

Impaired cross talk between keratinocytes (KCs) and immune cells is believed to contribute to the pathogenesis of chronic inflammatory skin diseases, such as psoriasis. We have previously identified KCs as a rich source of the serpin protease inhibitor vaspin (serpinA12), originally described as an adipokine in adipose tissue. Herein, we studied whether dysregulated vaspin expression in KCs contributes to the pathogenesis of psoriasis. We found vaspin expression to be closely associated to epidermal differentiation, with low levels in proliferating KCs and high levels in differentiated cells. Consistently, in human psoriasis and in a mouse model of a psoriasis-like skin inflammation, epidermal vaspin expression was significantly down-regulated. Down-regulation of vaspin in KCs resulted in decreased expression of differentiation-associated genes and up-regulation of interferon-inducible and inflammation-associated psoriasis signature genes. Vaspin was also shown to modulate the communication between KCs and inflammatory cells under co-culture conditions. A decrease in vaspin expression in KCs stimulated the secretion of tumor necrosis factor-α, IL-1β, IL-6, IL-8, and monocyte chemoattractant protein-1 by co-cultured dendritic cells, macrophages, monocytes, and neutrophils. Consequently, the application of vaspin inhibited myeloid cell infiltration in a mouse model of a psoriasis-like skin inflammation. In conclusion, vaspin expression by maturing KCs modulates cutaneous immune responses and may be involved in the pathogenesis of psoriasis.

Aeromonas veronii, a bacterium found in freshwater, is an unusual pathogen in healthy patients. We present a case report of a rare, aggressive subtype in a young, immunocompetent individual. History of injury in an aquatic environment and culture data are key for identification of the causal agent and should dictate acute clinical management and antibiotic therapy. Coverage should include cephalosporins, quinolones, or sulfas if Aeromonas is suspected, and adjusted depending on culture and sensitivity. Early surgical exploration, incision and drainage, and appropriate antimicrobial therapy are the cornerstones for successful treatment of these aggressive, sometimes life-threatening infections.

The understanding of subsurface events that cannot be directly observed is dependent on the ability to relate surface-based observations to subsurface processes. This is particularly important for nuclear explosion monitoring, as any future clandestine tests will likely be underground. We collected ground-based lidar and optical imagery using remote, very-low-altitude unmanned aerial system platforms, before and after several underground high explosive experiments. For the lidar collections, we used a terrestrial lidar scanner to obtain high-resolution point clouds and create digital elevation models (DEMs). For the imagery collections, we used structure-from-motion photogrammetry techniques and a dense grid of surveyed ground control points to create high-resolution DEMs. Comparisons between the pre- and post-experiment DEMs indicate changes in surface topography that vary between explosive experiments with varying yield and depth parameters. Our work shows that the relationship between explosive yield and the extent of observable surface change differs from the standard scaled-depth-of-burial model. This suggests that the surface morphological change from underground high explosive experiments can help constrain the experiments' yield and depth, and may impact how such activities are monitored and verified.

We propose an algorithm for dynamic optimization problems with inequality path constraints. It solves a sequence of approximated problems where the path constraint is imposed on a finite number of points. Between adjacent points, an approximating polynomial of the constraint value is calculated and an additional constraint is imposed on the maximum value of this polynomial. We consider Taylor and Hermite polynomials. New points are added based on constraint violations or large approximations errors of the approximating polynomials. We prove finite convergence to a feasible point assuming: (i) the dynamic optimization problem has a Slater point, (ii) pointwise constraints are respected at each iteration. We compare the performance of the algorithm with the algorithm by Fu et at. (Automatica 62, 2015, p. 184–192) for three small case studies and an up-to-date industrial application where we calculate optimal feed rates for a semi-batch emulsion polymerization reactor. The results show that our proposed algorithm needs to solve fewer subproblems, i.e. fewer iterations, at the cost of more constraints, resulting in smaller CPU times.

Dietary management of 418 adult patients with galactosaemia (from 39 centres/12 countries) was compared. All centres advised lactose restriction, 6 restricted galactose from galactosides +/- fruits and vegetables and 12 offal. 38% (n = 15) relaxed diet by: 1) allowing traces of lactose in manufactured foods (n = 13) or 2) giving fruits, vegetables and galactosides (n = 2). Only 15% (n = 6) calculated dietary galactose. 32% of patients were lost to dietetic follow-up. In adult galactosaemia, there is limited diet relaxation. (C) 2015 Elsevier Inc. All rights reserved.

Retinoic acid (RA) is required for development and homeostasis of the normal mammalian brain and may play a role in the initiation and progression of malignant brain tumors, such as the glioblastoma multiforme (GBM) and the gliosarcoma (Gsarc). The subpopulation of stem-like glioma cells (SLGCs) was shown to resist standard glioma radio-/chemotherapy and to propagate tumor regrowth. We used phenotypically distinct, self-renewing SLGC lines from six human GBMs, two Gsarcs, and two subcloned SLGC derivatives in order to investigate their responsiveness to all-trans retinoic acid (atRA) and to identify the RA-receptor (RAR) isotypes involved. In general, atRA exerted a pro-proliferative and pro-survival effect on SLGCs, though the efficacy was distinct. By means of RAR isotype-selective retinoids we disclosed that these effects were mediated by RARα and RARγ, except for one SLGC line, in which the pro-proliferative signal was induced by the RARβ-selective retinoid. Only one GBM-derived cell line (T1338) and a subpopulation of another (T1389) displayed neural differentiation in response to atRA. Differentiation of T1338 was induced by RARα and RARγ isotype-selective retinoids, associated with down-regulation of Sox2, and the failure to induce orthotopic tumors in the brains of SCID mice. The differential responsiveness of the SLGC lines appeared unrelated to the expression of RARβ, as (i) atRA augmented RAR isotype mRNA expression and particularly rarβ mRNA in all SLGC lines, (ii) rarβ promoter hypomethylation in the SLGC lines was not related to differentiation and (iii) the induction of T1338 differentiation was by RARα- and RARγ-selective ligands.

Summary Background The World Health Organization (WHO) recently proposed a new definition of severe asthma to facilitate standardized characterization of patients, and enable more accurate estimations of the prevalence of severe asthma. The aim of this study was to estimate the prevalence of severe asthma according to the WHO definition in children aged 12 years, in Stockholm, Sweden. Methods The birth cohort BAMSE enrolled 4089 children during 1994–96. Parental questionnaires provided information on asthma-related symptoms, diagnosis and medication from 3015 enrolled children at the age of 12 years. Severe asthma was defined as the presence of asthma, as well as continuous treatment with inhaled corticosteroids and long-acting beta-2 agonists, based on information from the Swedish prescribed drug register demonstrating prescriptions of at least 800 μg budesonide daily (or equivalent). Results The prevalence of asthma was 11% among 12-year-olds ( n = 329). Based on information from the Swedish prescribed drug register, seven children with asthma fulfilled the definition of severe asthma. The estimated prevalence corresponds to 0.23% (95% CI, 0.06–0.4) of the population, or 2.1% (95% CI, 0.5–3.7) of children with asthma. Based on assessed markers of asthma control, 3/7 with severe asthma were considered to have controlled asthma and 4/7 had partly or uncontrolled asthma. Conclusion Severe asthma appears rare both among 12-year-old schoolchildren with asthma and in the general population. Combining self-reported information from a population-based birth cohort with data from a drug register seems trustworthy in estimating severe asthma as defined by the WHO.

The effects of a triple combination of siRNAs targeting key scarring genes were assessed using an ex vivo organ culture model of excimer ablated rabbit corneas. The central 6 mm diameter region of fresh rabbit globes was ablated to a depth of 155 microns with an excimer laser. Corneas were excised, cultured at the air–liquid interface in defined culture medium supplemented with transforming growth factor beta 1 (TGFB1), and treated with either 1% prednisolone acetate or with 22.5 μM cationic nanoparticles complexed with a triple combination of siRNAs (NP-siRNA) targeting TGFB1, TGFB Receptor (TGFBR2) and connective tissue growth factor (CTGF). Scar formation was measured using image analysis of digital images and levels of smooth muscle actin (SMA) were assessed in ablated region of corneas using qRT-PCR and immunostaining. Ex vivo cultured corneas developed intense haze-like scar in the wounded areas and levels of mRNAs for pro-fibrotic genes were significantly elevated 3–8 fold in wounded tissue compared to unablated corneas. Treatment with NP-siRNA or steroid significantly reduced quantitative haze levels by 55% and 68%, respectively, and reduced SMA mRNA and immunohistostaining. This ex vivo corneal culture system reproduced key molecular patterns of corneal scarring and haze formation generated in rabbits. Treatment with NP-siRNAs targeting key scarring genes or an anti-inflammatory steroid reduced corneal haze and SMA mRNA and protein.

Background Wound care is the cornerstone of treatment for patients with epidermolysis bullosa (EB); however, there are currently no guidelines to help practitioners care for these patients. Objectives The objective of this study was to generate a list of recommendations that will enable practitioners to better care for patients with EB. Methods An expert panel generated a list of recommendations based on the best evidence available. The recommendations were translated into a survey, and sent to other EB experts to generate consensus using an online-based modified Delphi method. The list was refined and grouped into themes and specific recommendations. Results There were 15 respondents (45% response rate), with significant experience in the EB field (>10 years [67%]). Respondents included physicians (67%), nurses (17%), and allied health professionals (7%). There was more than 85% agreement for all the proposed items. These were further refined and grouped into 5 main themes (assessment and management of factors that impair healing, patient-centered concerns, local wound care, development of an individualized care plan, and organizational support) and 17 specific recommendations. Limitations There is a paucity of scientific evidence with most recommendations based on expert opinion. Conclusions These recommendations will provide practitioners with a framework for caring for these patients. Additional scientific research including effectiveness studies for everyday practice and expert consensus, may further refine these recommendations.

The physiological significance of the human vomeronasal duct (VND) is still unclear. The aim of the present study was to investigate the question whether mucosal responses obtained from the VND are different from those obtained from the respiratory epithelium. There were 15 healthy subjects (8 male, 7 female; age range 19-45 years; 14 normosmic subjects, 1 anosmic subject). All subjects participated in two sessions whereby the first session was used to acquaint them with the experimental conditions. For chemical stimulation, an olfactometer was used which delivered chemical stimulants without altering mechanical or thermal conditions at the stimulated nasal mucosa. For stimulation we used substances previously reported to produce vomeronasal activation ("estra"=estra-1,3,5(10),16-tetraen-3ol and "andro"=androsta-4,16-dien-3-on); in addition, gaseous CO(2) was used as a non-odorous, relatively specific stimulant of the trigeminal nerve. Placement of electrodes either in the VND or on the respiratory epithelium was performed under endoscopical guidance. Subjects rated the overall intensity of the stimuli, the strength of trigeminally mediated sensations, and the hedonic tone of the stimulants. Responses could not be recorded from all subjects. For the remaining 7 subjects, intensity was strongest for CO(2) stimuli (p

Resume Introduction La sarcoidose est une maladie granulomateuse, sans necrose caseeuse, d’etiologie inconnue. Etat des connaissances La sarcoidose des voies aerodigestives superieures (SVAS) regroupe la sarcoidose nasosinusienne (SNS) et la sarcoidose pharyngolaryngee (SPL) qui peuvent etre isolees ou s’integrer dans une sarcoidose multisystemique. La symptomatologie clinique est proteiforme, aspecifique. L’evolution et l’histoire naturelle sont mal connues et imprevisibles. Le diagnostic reste difficile. Le traitement n’est pas standardise pour un pronostic a long terme souvent decevant. Perspectives Face a ces incertitudes diagnostiques et therapeutiques, les auteurs cherchent a realiser un document de synthese a partir de leur experience, de leurs publications anterieures et des donnees de la litterature et a evaluer l’interet de la tomographie par emission de positons couplee a la tomodensitometrie (TEP TDM) au 18F–Fluorodesoxyglucose (18F-FDG) dans la prise en charge de la SVAS. Conclusions Si la biopsie orientee reste l’etalon d’or pour confirmer le diagnostic de SVAS, l’imagerie joue un role important : TDM permettant de decrire et de classer la SNS en deux stades bien correles avec la severite et le profil evolutif ; TEP TDM au 18F-FDG permettant une cartographie morphofonctionnelle des lesions sarcoidosiques actives, tant pour le diagnostic que pour le suivi therapeutique.

There are fundamental differences between acute wounds that proceed to uncomplicated healing and those that become chronic wounds. Non-healing or chronic wounds can result from a combination of overlapping factors that prevent healing, including local tissue ischemia, repetitive trauma and ischemia/reperfusion injury, presence of tissue necrosis, impaired cellular and systemic host response to stress, and critical bacterial contamination. The bacterial burden in the wound contributes to a sustained inflammatory state, which inhibits normal progression to the proliferative phase of healing, thereby preventing restoration of tissue integrity. Appropriate wound bed preparation removes local barriers to healing and optimizes the tissue environment to achieve wound healing. It is an essential element of wound management that advances endogenous healing as well as the efficacy of topical and other wound therapy. This article will summarize a systematic approach to wound bed preparation using the "TIME" principle, and will highlight important advances in topical wound care.

Corneal ulcers can cause significant loss of vision from scarring and astigmatism, but rapid management can limit the destruction and improve outcomes. Infectious ulcers usually resolve with antimicrobial treatment. Noninfectious ulcers, however, present a diagnostic and therapeutic challenge. They can often be resolved by eliminating toxic medications and providing surface support with lubrication and collagenase inhibitors, but resistant ulcers may need more aggressive therapy with bandage contact lenses, tarsorrhaphy, or autologous serum. Ulcers impending perforation require urgent surgical management (e.g., tissue glue, conjunctival flaps, or keratoplasty). Topical steroids are useful when the ulceration is secondary to inflammatory mediators, but they are contraindicated in corneal melts with minimal inflammation, such as those associated with Sjogren syndrome. Systemic immunomodulation is required in addition to topical therapy in the presence of autoimmune disease. Understanding of the pathological processes that occur in different types of corneal ulcers is essential to formulation of a logical and effective treatment plan. Newer, more targeted treatment modalities may offer treatment options that have greater efficacy and fewer adverse effects.

Four of the 5 biotin-dependent carboxylases (BDC) are in the mitochondria. BDC replace intermediates in the Krebs [tricarboxylic acid (TCA)] cycle that are regularly removed for the synthesis of key metabolites such as heme or amino acids. Heme, unlike amino acids, is not recycled to regenerate these intermediates, is not utilized from the diet, and must be synthesized in situ. We studied whether biotin deficiency (BD) lowers heme synthesis and whether mitochondria would be disrupted. Biotin-deficient medium was prepared by using bovine serum stripped of biotin with charcoal/dextran or avidin. Biotin-deficient primary human lung fibroblasts (IMR90) lost their BDC and senesced before biotin-sufficient cells. BD caused heme deficiency; there was a decrease in heme content and heme synthesis, and biotin-deficient cells selectively lost mitochondrial complex IV, which contains heme-a. Loss of complex IV, which is part of the electron transport chain, triggered oxidant release and oxidative damage, hallmarks of heme deficiency. Restoring biotin to the biotin-deficient medium prevented the above changes. Old cells were more susceptible to biotin shortage than young cells. These findings highlight the biochemical connection among biotin, heme, and iron metabolism, and the mitochondria, due to the role of biotin in maintaining the biochemical integrity of the TCA cycle. The findings are discussed in relation to aging and birth defects in humans.

Multipotent neural stem cells have been isolated from the adult [Kirschenbaum B, Nedergaard M, Preuss A, Barami K, Fraser RA, Goldman SA. In vitro neuronal production and differentiation by precursor cells derived from the adult human forebrain. Cereb Cortex 1994;4(6):576-89; Laywell ED, Kukekov VG, Steindler DA. Multipotent neurospheres can be derived from forebrain subependymal zone and spinal cord of adult mice after protracted postmortem intervals. Exp Neurol 1999;156:430-3; Pluchino S, Quattrini A, Brambilla E, Gritti A, Salani G, Dina G, et al. Injection of adult neurospheres induces recovery in a chronic model of multiple sclerosis. Nature 2003;422:688-94] and embryonic [Vescovi AL, Parati EA, Gritti A, Poulin P, Ferrario M, Wanke E, et al. Isolation and cloning of multipotential stem cells from the embryonic human CNS and establishment of transplantable human neural stem cell lines by epigenetic stimulation. Exp Neurol 1999;156:71-83] central nervous system (CNS). In addition, neural cells can be obtained from sources other than the CNS by differentiating stem cells from a non-neural source down a neural lineage. This has previously been performed with pluripotent embryonic stem cells and adult stem cells derived from rat bone marrow [Woodbury D, Schwarz EJ, Prockop DJ, Black IB. Adult rat and human bone marrow stromal cells differentiate into neurons. J Neurosci Res 2000;61:364-70; Woodbury D, Reynolds K, Black IB. Adult bone marrow stromal stem cells express germline, ectodermal, endodermal, and mesodermal genes prior to neurogenesis. J Neurosci 2002;69(6):908-17] and skeletal muscle [Romero-Ramos M, Vourc'h P, Young HE, Lucas PA, Wu Y, Chivatakarn O, et al. Neuronal differentiation of stem cells isolated from adult muscle. J Neurosci Res 2002;69:894-907]. Previously, we have isolated adult stem cells from human skeletal muscle with the potential to differentiate into mesoderm, ectoderm, and endoderm. The following in vitro experiments were designed to determine whether human adult stem cells behaved similarly to rat adult stem cells when both were isolated from skeletal muscle by the same procedure [Romero-Ramos M, Vourc'h P, Young HE, Lucas PA, Wu Y, Chivatakarn O, et al. Neuronal differentiation of stem cells isolated from adult muscle. J Neurosci Res 2002;69:894-907] and subjected to the same protocols to induce neurogenesis. The neural phenotypes that were created through the neurococktail or neurosphere protocol were analyzed for neural characteristics through morphology and immunohistochemistry antibody labeling for proteins to neurons (RT-97, beta-tubulin III, NF-160, NF-200, and synapsin), oligodendrocytes (CNPase and RIP), and astrocytes (GFAP). A calcium uptake assay also showed response to the neuronal excitotoxic agent glutamic acid. In conclusion, the neural differentiated stem cells derived from adult skeletal muscle may be a less invasive alternative for the treatment of CNS disorders over CNS derived neural stem cells.

A marked difference in spin relaxation function was found between positive muon in oxy-hemoglobin containing human blood versus that in deoxy-hemoglobin containing human blood, at room temperature and under zero and low external longitudinal magnetic field. Future extension to the functional study of the human brain is suggested.

Spectroscopic (IR, 31P NMR and UV–Vis) and electrochemical studies on fac-[Mn(CO)3(L′–L′)(L)]0/+,where L′–L′ = 1,2-bis(diphenylphosphino)ethane (dppe) or 1,10-phenanthroline (phen) and L = bromide, triflate, imidazole (im), isonicotinamide (isn) or N-(2-hydroxyethyl)isonicotinamide (heisn), were undertaken to understand the effect of various ligands on the CO–Mn–L and CO–Mn–(L′–L′) bonding characteristics of these complexes. Crystal structures for L = triflate/L′–L′ = dppe, L = triflate/L′–L′ = phen and L = isn/L′–L′ = phen are reported and they show that the two Mn–O(OSO2CF3) and Mn–N(isn) distances are similar. The tricarbonyl complexes exhibit two major bands in the 250–300 and 350–450 nm region of the UV–Vis spectrum. The lowest energy bands have been assigned as a contribution from both the metal-centered (MC) and metal to ligand (dπ → L′–L′) charge transfer (MLCT) transitions. The energy of this maximum absorption decreases in the order Br− ∼ triflate > im > isn ∼ heisn. The cyclic four-component mechanism fac - Mn ( I ) → - e fac - Mn ( II ) → mer - Mn ( II ) → + e mer - Mn ( I ) → fac - Mn ( I ) was observed at room temperature by voltammetric techniques for all the cases. On the basis of d metal orbital splitting, an electronic molecular orbital diagram is proposed. In this model, the ligands along the z-axis play a relevant role in the reverse of the HOMO energies of the fac/mer isomers by stabilizing the metal d z 2 orbital relative to dxy in mer-Mn(II).

In this study, moisture, ash, amylose, phosphorous content, and the gelatinization profiles of starches isolated from Colocasia esculenta (taro), and Xanthosoma sagitifolium (tannia) storage organs were evaluated. The gelatinization profile and the changes in the heat flow or enthalpy during the gelatinization process were evaluated by DSC methodology. The phosphorous and amylose content were also analyzed by a colorimetric method. The results show that the amylose content of the starch isolated from Xanthosoma sagittifolium is higher than those shown by Colocasia esculenta and Manihot esculenta Crantz starches. The phosphorous content was higher in Xanthosoma sagittifolium than Colocasia esculenta or the commercial Manihot esculenta C. starches. The gelatinization profile range is wider in Manihot esculenta C. than the other two starches. Differences in these parameters may affect the functional properties of the products formulated with these starches. The most significant relationship between parameters was found between the amylose and gelatinization profile and enthalpic change and ash.

Glucose binding protein (GBP) from Escherichia coli has been widely used to develop minimally invasive glucose biosensors for diabetics. To develop a cell-based glucose biosensor, it is essential to functionally display GBP on the cell surface. In this study, we designed a molecular structure to display GBP on the outer membrane of E. coli . We fused GBP with the first nine N-terminal residues of Lpp (major E. coli lipoprotein) and the 46–150 residues of OmpA (an outer membrane protein of E. coli ). With this molecular design, we have successfully displayed GBP on the surface of E. coli . Using FITC-conjugated Dextran, we demonstrated that glucose’s binding sites of surface-displayed GBP were accessible to glucose. Furthermore, we showed that glucose transport in a GBP-deficient E. coli NM303 could be restored by displaying GBP on the surface of NM303. 0.51 h −1 of specific growth rate was attained for NM303/pESDG grown in M9 minimal medium supplemented with 2 g/l glucose, whereas no growth was observed for NM303 in the same medium. Both NM303 and NM303/pESDG grew in M9 medium supplemented with 1 mM of fucose. Because cell surface is an interface between intracellular and extracellular molecular events, this technique paves a way to develop cell-based glucose biosensors.

Peptide deformylases (PDFs) catalyze the removal of the N-terminal formyl group from nascent polypeptides. In spite of the vast amount of literature on PDF, no information whatsoever is available on its use in organic synthesis. To be able to explore the potential of E. coli PDF (EcPDF) in biocatalytic applications, a simple and efficient purification procedure was developed. This method, which was based on one affinity chromatographic step, furnished about 200 mg of pure EcPDF from 1 L of E. coli culture. The enzyme combined a good activity (tof ≥5 s−1) with an almost complete enantioselectivity (E ratio >500) in the resolution of N-formylated α- and β-amino acids, α-amino acid amides and α-aminonitriles. N-Formyl derivatives of non-functionalized amines and β-amino alcohols were hydrolyzed with low to moderate activity and enantioselectivity. EcPDF was also successfully applied in the enantioselective formylation of α-aminonitriles, yielding, e.g. (S)-N-formyl-phenylalanine nitrile with >99.5% ee. The enzyme also proved very suitable for the mild and selective deprotection of N-formyl peptides as was shown for N-formyl-Leu-Tle-NHCH3. This deprotection increased the diastereomeric excess of the dipeptide, which was unsatisfactory because of racemization of the N-terminal amino acid in the chemical peptide coupling step.

Purpose. Excessive scarring following trauma or surgery of cornea, conjunctiva or retina can greatly impair visual outcome. At present, no agents are clinically available that selectively reduce activity of genes that regulate fibrosis. Connective tissue growth factor (CTGF) has been linked to fibrosis in several tissues, including cornea and conjunctiva. In this study, hammerhead ribozymes targeting CTGF mRNA were synthesized, kinetic parameters were measured, and the effect on TGF-β-mediated cell proliferation was measured in cultured human fibroblasts. Methods. The mRNA sequence of human CTGF was scanned for potential hammerhead ribozyme cleavage sites, and predicted secondary folding structures around the sites were calculated. Synthetic 12mer ribozymes and 33mer oligonucleotide mRNA targets corresponding to two sites were synthesized, and kinetic constants calculated from Hanes–Wolff plots of in vitro cleavage reactions. The ribozyme with higher percentage cleavage and kinetic rate was cloned into an expression plasmid (pTR-UF21) and stably transfected into cultured human fibroblasts. An inactive ribozyme plasmid served as a negative control. The effects of the ribozyme on expression of TGF-β-induced CTGF mRNA and protein levels were measured using ELISA and real-time TaqMan quantitative RT-PCR. Finally, the effect of the CTGF ribozyme on TGF-β-mediated proliferation of fibroblasts was measured using a non-radioactive cell proliferation microtiter assay. Results. Of the eight potential hammerhead ribozyme cleavage sites in human CTGF mRNA, two sites (CHR 745, and CHR 859) were identified with optimal secondary folding. CHR 859 cleaved 94% of the target mRNA, compared to 46% cleavage for CHR 745 after 16 hr of reaction. CHR 859 had a Km of 1·56 μ m and a Kcat of 2·97 min−1, while CHR 745 had a Km of 7·80 μ m and a Kcat of 5·7 min−1. The turnover numbers (Kcat/Km) of CHR 859 and CHR 745 were 1·9×106 m min−1 and 7·4×105 m min−1, respectively, indicating CHR 859 is 2·6 times more efficient than CHR 745 in destroying CTGF mRNA. Stable transfection of CHR 859 into human fibroblasts reduced CTGF mRNA levels 55% and protein levels 72% compared to the inactive ribozyme control. Furthermore, TGF-β-induced cell proliferation was reduced 90% in fibroblasts stably transfected with CHR 859 compared to control cell groups. Conclusions. The CHR 859 hammerhead ribozyme cleaved human CTGF mRNA with high kinetic efficiency in vitro, effectively reduced levels of CTGF mRNA and protein in cultured human fibroblasts, and blocked TGF-β-induced cell proliferation without nonspecific toxicity. These data support the concept that CTGF mediates TGF-β-induced cell proliferation, and imply that regulating CTGF expression with ribozymes may be effective in reducing ocular scarring.

Excessive cell-mediated tissue contraction after injury can lead to morbid contractile scarring in the body. In the eye this can cause blindness because of posterior capsule opacification, proliferative vitroretinopathy, failure of glaucoma filtration surgery, and corneal haze. During repair, transforming growth factor (TGF)-β and connective tissue growth factor (CTGF) genes are co-ordinately expressed. Although TGF-β and CTGF stimulate new matrix deposition, their role and regulation during contractile scarring is unknown. In this study, an in vitro model of collagen matrix contraction culminating from tractional forces generated by fibroblasts showed that both TGF-β1 and CTGF-stimulated contraction. Using a specific anti-sense oligodeoxynucleotide to CTGF the procontractile activity of TGF-β1 was found to be mediated by CTGF. During contraction fibroblasts produced similar levels of matrix metalloproteinases (MMPs)-2 and -9 with TGF-β1 or CTGF and a modest increase in MMP-1 with CTGF only (indicated by zymography and enzyme-linked immunosorbent assay). The requirement of MMPs for contraction was demonstrated using a broad-spectrum synthetic inhibitor. This study demonstrates a new function for CTGF in mediating matrix contraction by fibroblasts involving MMPs and suggests a novel regulatory mechanism for TGF-β-stimulated contraction. Inhibition of CTGF activity or gene transcription could be a suitable target for anti-scarring therapies.

Dendritic cells (DC) based vaccinations have been widely used for the induction of anti-tumoral immunity in clinical studies. Antigen loading of DC with whole tumor cell preparations is an attractive method whenever tumor cell material is available. In order to determine parameters for the loading procedure, we performed dose finding and timing experiments. We found that apoptotic and necrotic melanoma cells up to a ratio of one-to-one, equivalent to 1mg/ml protein per 1 � 10 6 DC, can be added to monocyte derived DC without effecting DC recovery extensively. Using the isolated protein content of tumor cells (lysate) as a parameter, up to 5 mg/ml protein per 1 � 10 6 DC can be added. To achieve significant protein uptake at least 1 mg/ml of protein have to be added for more than 24 h as tested with FITC-labelled ovalbumin. Maturation inducing cytokines can be added simultaneously with the tumor cell preparations to immature DC without affecting the uptake. Furthermore, we tested the feasibility of cryopreservation of loaded and matured DC to facilitate the generation of ready to use aliquots. DC were cryopreserved in a mix of human serum albumin, DMSO and 5% glucose. After thawing, surface expression of molecules indicating the mature status (CD83, costimulatory and MHC molecules), was found to be unaltered. Furthermore, cryopreserved DC kept the capability to stimulate allogenic T-cell proliferation in mixed leukocyte reactions at full level. Loaded and matured DC pulsed with influenza matrix peptide (IMP) retained the capacity to induce the generation of IMP-specific cytotoxic Tlymphocytes after cryopreservation as measured by ELISPOT and tetramer staining. The expression of the chemokine receptor CXCR-4 and CCR-7 remained unaltered during cryopreservation and the migratory responsiveness towards MIP-3h was unaltered as measured in a migration assay. Thus we conclude that the large scale loading and maturation of DC with whole tumor cell preparations can be performed in a single session. These data will facilitate the clinical application of DC loaded with whole tumor cell preparations. D 2003 Elsevier Science B.V. All rights reserved.

Healing of corneal injuries is an exceptionally complex process involving the integrated actions of multiple growth factors, cytokines, and proteases produced by epithelial cells, stromal keratocytes, inflammatory cells, and lacrimal gland cells. Following corneal injury, basal epithelial cells migrate and proliferate in response to chemotactic cytokines and mitogenic growth factors, including epidermal growth factor and keratinocyte growth factor. Simultaneously, keratocytes adjacent to the injured area undergo apoptosis under the Fas/Fas ligand system, while more distant keratocytes transform into activated fibroblasts and migrate into the wound, where they begin synthesizing new extracellular matrix components that form the scar tissue under the dominant influence of the TGFβ/CTGF system. Epithelial cells and activated stromal fibroblasts also secrete growth factors and cytokines that have paracrine and autocrine functions. Corneal repair proceeds for the next several weeks to months, during which time the gene expression profile slowly returns to the pre-injury pattern and the provisional scar matrix slowly remodels by actions of matrix metalloproteinases. While minor epithelial injuries heal by regeneration of normal architecture, large stromal injuries heal by repair with irregular scar tissue that impairs the optical properties of the cornea. Also, if the integrated regulation of the wound healing process is interrupted at any point, the wound fails to heal properly and a corneal ulcer develops. Better understanding of the cellular and molecular changes that occur during repair of corneal wounds will provide the opportunity to design agents that selectively modulate key phases of corneal wound healing, resulting in scars that more closely resemble normal corneal architecture.

The transforming growth factor betas are of major importance in the wound repair process; however, no studies to date have investigated the role of the transforming growth factor beta receptors in chronic venous leg ulcers or what effect healing has on these proteins. To determine whether the transforming growth factor beta peptides and their receptors are expressed in chronic venous wounds, we used immunofluorescent analysis and quantitative competitive reverse transcription polymerase chain reaction to identify the protein and mRNA expression, respectively. Biopsy samples from wounds and normal skin were collected from 12 patients with chronic venous leg ulcers and three patients undergoing reconstructive surgery, respectively. Additionally four of the chronic venous leg ulcer patients were re-biopsied between 2 and 8 wk after the first biopsy when the wounds had entered the healing phase. The tissue excised from the ulcers included the surrounding intact skin, the ulcer edge, and the ulcer base. Immunofluorescent staining for transforming growth factors beta1, beta2, and beta3 was observed within the epidermis of the skin surrounding the chronic venous ulcers and in fibroblasts and inflammatory cells of the dermis, although this staining was not as strong as that seen in normal unwounded skin. Very little staining could be seen within the ulcers for any of the ligands, however. In contrast the transforming growth factor beta type I receptor was observed throughout the ulcers and the normal unwounded skin biopsies, particularly in the basal epidermal cells. No immunofluorescence for the type II transforming growth factor beta receptor was observed in any of the ulcer biopsies investigated, although it was observed throughout the epidermis and in fibroblasts and inflammatory cells in the surrounding skin. Quantitative, competitive reverse transcription polymerase chain reaction was used to analyze mRNA expression for transforming growth factor beta1 and the type II receptor in the nonhealing ulcers and normal unwounded skin biopsies. These studies revealed that transforming growth factor beta1 and transforming growth factor beta receptor II mRNA was expressed in all the chronic nonhealing ulcers albeit at very low levels for the type II receptor. In marked contrast to the staining observed in nonhealing chronic ulcers, positive immunostaining was observed for the transforming growth factor betas and both the type I and type II receptors in healing ulcers. These results suggest that the absence of a viable receptor complex for the transforming growth factor betas in nonhealing chronic venous ulcers may contribute to wound chronicity.

Telomerase is a reverse transcriptase responsible for adding simple sequence repeats to chromosome 3'-ends. The template for telomeric repeat synthesis is carried within the RNA component of the telomerase ribonucleoprotein complex. Telomerases can copy their internal templates with repeat addition processivity, reusing the same template multiple times in the extension of a single primer. For some telomerases, optimal repeat addition processivity requires high micromolar dGTP concentrations, a much higher dGTP concentration than required for processive nucleotide addition within a repeat. We have investigated the requirements for dGTP-dependent repeat addition processivity using recombinant Tetrahymena telomerase. By altering the template sequence, we show that repeat addition processivity retains the same dGTP-dependence even if dGTP is not the first nucleotide incorporated in the second repeat. Furthermore, no dNTP other than dGTP can stimulate repeat addition processivity, even if it is the first nucleotide incorporated in the second repeat. Using structural variants of dGTP, we demonstrate that the stimulation of repeat addition processivity is specific for dGTP base and sugar constituents but requires only a single phosphate group. However, all nucleotides that stimulate repeat addition processivity also inhibit or compete with dGTP incorporation into product DNA. By assaying telomerase complexes reconstituted with a variety of altered templates, we find that repeat addition processivity has an unanticipated template or product sequence specificity. Finally, we show that a novel, nascent product DNA binding site establishes dGTP-dependent repeat addition processivity.

Objective: This study was undertaken to determine whether obstetric factors affect the concentrations of matrix metalloproteinase 9 and tissue inhibitor of metalloproteinase 1 in the amniotic fluid. Study Design: We prospectively collected amniotic fluid samples from 109 women at various stages of pregnancy and labor and determined matrix metalloproteinase 9 and tissue inhibitor of metalloproteinase 1 concentrations by means of enzyme-linked immunosorbent sandwich assay systems. With multiple regression analysis we evaluated relationships between amniotic fluid matrix metalloproteinase 9 concentration and tissue inhibitor of metalloproteinase 1 concentration and the following factors: gestational age, presence of labor, cervical dilatation, membrane status, presence of clinical chorioamnionitis, and microbial colonization of the amniotic fluid. Results: The detectable presence of amniotic fluid matrix metalloproteinase 9 was independently associated with intra-amniotic infection, labor, cervical dilatation, and spontaneous rupture of membranes. Chorioamnionitis and amniotic fluid matrix metalloproteinase 9 concentrations were correlated with tissue inhibitor of metalloproteinase 1 levels. Conclusions: Intra-amniotic infection, advanced labor, and rupture of membranes before the onset of labor were independently associated with the presence of matrix metalloproteinase 9 in the amniotic fluid. Both pathologic and physiologic processes appear to produce shifts in the balance between degradation and synthesis of the extracellular matrix. (Am J Obstet Gynecol 2001;184:159-64.)

The dissolution of highly viscous dispersions composed of complexes of Concanavalin A (Con A) and high molecular-weight dextran (2000 kDa) in response to glucose was studied by using surface plasmon resonance (SPR). The viscous property of Con A/dextran sols is based on intermolecular affinity bonds between dextran and Con A. These receptor/ligand cross-linkages can be reversibly cleaved by the competing analyte glucose, resulting in a sol with a strongly reduced viscosity. We tested the concept of using the kinetics of sol dissolution as an analytical tool for glucose determination. Under this premise, a thin layer of the Con A/dextran dispersion was placed on the gold surface of a novel SPR-probe. The sol's dissolution, when placed in contact with bulk solutions of various glucose concentrations, was monitored by measuring the shift of resonance angle. An increased rate of sol dissolution at increasing levels of glucose concentration was found, presumably due to a decrease of the sol viscosity which led to a faster diffusion and mass transport of the sol components into the bulk solution. The thickness of the sol layer turned out to be the rate-limiting step of the dissolution kinetics. Other factors such as sugar specificity and temperature had a significant affect on the kinetics of sol dissolution also. However, matrix effects of the test solution effected the overall kinetics of the solution only to a minimal extent, rendering the method interesting for biosensing purposes.

Laboratory experiments have demonstrated electroosmosis to be very effective for removing a water-soluble organic from low-permeability silty clay soil from a DuPont site in the eastern U.S. Concentrations were reduced below detection levels by electroosmotically driving 1.5 pore volumes of water through the soil. Neutralizing the fluid at the anode with lime or sodium hydroxide significantly increased the efficiency of the process. An in-situ field trial is under way. A simple economic analysis is presented, which estimates the cost of applying the technology for this application full-scale to be $78 m −3 ($63 per cubic yard) for a three-year remediation project.

Early morphogenesis of mouse submandibular glands begins on late day 11 of fetal development when the epithelium begins to bud from the surrounding mandibular mesenchyme. Using total RNA collected from fetal BALB/c submandibular glands, steady-state levels of mRNA expression for extracellular matrix molecules were measured using quantitative competitive reverse transcription-polymerase chain reaction (RT-PCR). By comparing the PCR amplification products of both the cellular mRNA and a synthetic template, pMATRIX, it was possible to measure the direct expression of collagens alpha2(I), alpha1(III), alpha1(IV), fibronectin, laminin B2, elastin and lysyl oxidase genes. There was an observed trend for an increasing concentration of collagen alpha2(I), collagen alpha1(III) and lysyl oxidase mRNA molecules per cell on day 16 of development. The relative abundance of elastin mRNA was detectable only on day 16. Fibronectin and laminin B2 were more constitutively present but had their highest copy number per cell on day 16. The presence of extracellular-matrix protein was confirmed by immunohistochemistry using day-16 fetal glands and adult glands. With the construction of the pMATRIX supertemplate and the advent of quantitative, competitive RT-PCR technology, it has been possible to measure small changes in the steady-state concentrations for extracellular-matrix mRNA during salivary gland development.