Decision-Making for HIV AIDS Prevention: Altruism and the Moral Norm.

The purpose of this enquiry was to understand how gay men form and maintain their attitudes toward HIV transmission preventative behaviors. Autobiographical life histories of sixteen gay men showed that once they acquired knowledge of preventative behavior they consistently adhered to that behavior. They adhered because of fear of HIV infection and because they held a moral norm that obligated them to behave altruistically (Schwartz, 1977) to protect not only themselves, but also their sex partners, loved ones, and their positive self-evaluation. They saw their HIV negative status, and their adherence, as pre-requisite and enabler for achieving their goals in life. Dick and Basu's (1994) Framework for Customer Loyalty, a commercial marketing communications theoretical framework, explains development and maintenance of these men's loyalty (their consistent adherence). This understanding, within a marketing communications framework, will inform development of social marketing communications aiming to increase adherence to behaviors that prevent HIV transmission.

Frugal Innovation: Enabling Mechanical Ventilation During Coronavirus Disease 2019 Pandemic in Resource-Limited Settings.

ICUs worldwide are facing resource shortages including increased need for provision of invasive mechanical ventilation during the current coronavirus disease 2019 pandemic. Fearing shortage of ventilators, many private companies and public institutions have focused on building new inexpensive, open-source ventilators. However, designing and building new ventilators is not sufficient for addressing invasive mechanical ventilation needs in resource-limited settings. In this commentary, we highlight additional interdependent constraints that should be considered and provide a framework for addressing these constraints to ensure that the increasing stockpile of open-source ventilators are easily deployable and sustainable for use in resource-limited settings.

An Integrated Circuit for Chip-Based Analysis of Enzyme Kinetics and Metabolite Quantification.

We have created a novel chip-based diagnostic tools based upon quantification of metabolites using enzymes specific for their chemical conversion. Using this device we show for the first time that a solid-state circuit can be used to measure enzyme kinetics and calculate the Michaelis-Menten constant. Substrate concentration dependency of enzyme reaction rates is central to this aim. Ion-sensitive field effect transistors (ISFET) are excellent transducers for biosensing applications that are reliant upon enzyme assays, especially since they can be fabricated using mainstream microelectronics technology to ensure low unit cost, mass-manufacture, scaling to make many sensors and straightforward miniaturisation for use in point-of-care devices. Here, we describe an integrated ISFET array comprising 2(16) sensors. The device was fabricated with a complementary metal oxide semiconductor (CMOS) process. Unlike traditional CMOS ISFET sensors that use the Si3N4 passivation of the foundry for ion detection, the device reported here was processed with a layer of Ta2O5 that increased the detection sensitivity to 45 mV/pH unit at the sensor readout. The drift was reduced to 0.8 mV/hour with a linear pH response between pH 2-12. A high-speed instrumentation system capable of acquiring nearly 500 fps was developed to stream out the data. The device was then used to measure glucose concentration through the activity of hexokinase in the range of 0.05 mM-231 mM, encompassing glucose's physiological range in blood. Localised and temporal enzyme kinetics of hexokinase was studied in detail. These results present a roadmap towards a viable personal metabolome machine.

Lymphadenopathy: defining a palpable lymph node.

BACKGROUND: The threshold size required to detect lymphadenopathy via palpation has never been formally determined. The purpose of this study was to determine the threshold, sensitivity, and error of node palpation and how this changes with experience. METHODS: Lymphadenopathy models were created using polyvinyl alcohol cryogel (PVA-C) to mimic tissue tactility. Node diameter ranged from 0.5 to 4 cm. Study subjects were medical students, otolaryngology residents, and otolaryngology consultants. Each subject provided 22 estimates of size. Primary outcomes were the sensitivity, error (true vs estimated size), and threshold of palpation. RESULTS: Thirty subjects completed the study. Sensitivity was 60%, 74%, and 86% for students, residents, and consultants, respectively (p < .01). Error was 0.88 cm, 0.61 cm, and 0.57 cm, respectively (p < .05). Palpation threshold was 1.32 cm, 0.83 cm, and 0.75 cm, respectively (p < .05). All participants detected nodes ≥2 cm, whereas consultants detected nodes ≥1 cm. CONCLUSION: Experience is associated with decreased palpation threshold and error, and increased sensitivity. Educational interventions should target nodes <2 cm.

Smooth muscle phenotypic modulation--a personal experience.

The idea that smooth muscle cells can exist in multiple phenotypic states depending on the functional demands placed upon them has been around for >5 decades. However, much of the literature today refers to only recent articles, giving the impression that it is a new idea. At the same time, the current trend is to delve deeper and deeper into transcriptional regulation of smooth muscle genes, and much of the work describing the change in biology of the cells in the different phenotypic states does not appear to be known. This loss of historical perspective regarding the biology of smooth muscle phenotypic modulation is what the current article has tried to mitigate.

Development of a tissue-mimicking neck model for medical education.

BACKGROUND: The ability to palpate neck masses and lymph nodes and appreciate qualities such as size, location, and consistency is critical for patient care and an important clinical skill for all physicians. Medical students currently learn neck palpation by practicing on healthy, standardized patients; however, studies of similar procedures have shown that educational models with simulated pathology help improve technique and confidence. OBJECTIVE: Our goal was to create a tissue-mimicking neck model with palpable masses. METHODS: Iterative design and development of a high-fidelity neck model using polyvinyl alcohol-cryogel (PVA-C), a nontoxic and biocompatible polymer that exhibits favourable tissue-mimicking elastic properties. Model geometries were digitally reconstructed from high-resolution cadaveric magnetic resonance imaging sections to create physical moulds through stereolithography. PVA-C was formulated to mimic the characteristics of human tissue. RESULTS: A life-like neck model was built and consists of these components: muscle, larynx, spine, soft tissue, pathologic nodes, and skin. DISCUSSION: A final neck model prototype has been completed and will be evaluated by otolaryngology consultants and residents for face and construct validity and assessed in a randomized, controlled trial to evaluate how it impacts students' ability to detect neck masses.

Contribution of inter-subunit interactions to the thermostability of Pyrococcus furiosus citrate synthase.

Using citrate synthase from the hyperthermophile Pyrococcus furiosus (PfCS) as our test molecule, we show through guanidine hydrochloride-induced unfolding that the dimer separates into folded, but inactive, monomers before individual subunit unfolding takes place. Given that forces across the dimer interface are vital for thermostability, a robust computational method was derived that uses the University of Houston Brownian Dynamics (UHBD) program to calculate both the hydrophobic and electrostatic contribution to the dimerisation energy at 100°C. The results from computational and experimental determination of the lowered stability of interface mutants were correlated, being both of the same order of magnitude and placing the mutant proteins in the same order of stability. This computational method, optimised for hyperthermophilic molecules and tested in the laboratory, after further testing on other examples, could be of widespread use in the prediction of thermostabilising mutations in other oligomeric proteins for which dissociation is the first step in unfolding.

Transcription factors expressed in olfactory bulb local progenitor cells revealed by genome-wide transcriptome profiling.

The local progenitor population in the olfactory bulb (OB) gives rise to mitral and tufted projection neurons during embryonic development. In contrast, OB interneurons are derived from sources outside the bulb where neurogenesis continues throughout life. While many of the genes involved in OB interneuron development have been characterized, the genetic pathways driving local progenitor cell differentiation in this tissue are largely unknown. To better understand this process, we used transcriptional profiling to monitor gene expression of whole OB at daily intervals from embryonic day 11 through birth, generating a compendium of gene expression encompassing the major developmental events of this tissue. Through hierarchical clustering, bioinformatics analysis, and validation by RNA in situ hybridizations, we identified a large number of transcription factors, DNA binding proteins, and cell cycle-related genes expressed by the local neural progenitor cells (NPCs) of the embryonic OB. Further in silico analysis of transcription factor binding sites identified an enrichment of genes regulated by the E2F-Rb pathway among those expressed in the local NPC population. Together these results provide initial insights into the molecular identity of the OB local NPC population and the transcription factor networks that may regulate their function.

Discovery of a Novel Series of CRTH2 (DP2) Receptor Antagonists Devoid of Carboxylic Acids.

Antagonism of the CRTH2 receptor represents a very attractive target for a variety of allergic diseases. Most CRTH2 antagonists known to date possess a carboxylic acid moiety, which is essential for binding. However, potential acid metabolites O-acyl glucuronides might be linked to idiosynchratic toxicity in humans. In this communication, we describe a new series of compounds that lack the carboxylic acid moiety. Compounds with high affinity (K i < 10 nM) for the receptor have been identified. Subsequent optimization succeeded in reducing the high metabolic clearance of the first compounds in human and rat liver microsomes. At the same time, inhibition of the CYP isoforms was optimized, giving rise to stable compounds with an acceptable CYP inhibition profile (IC50 CYP2C9 and 2C19 > 1 µM). Taken together, these data show that compounds devoid of carboxylic acid groups could represent an interesting alternative to current CRTH2 antagonists in development.

Optimization of the Central Core of Indolinone-Acetic Acid-Based CRTH2 (DP2) Receptor Antagonists.

New spiroindolinone antagonists of CRTH2 are described. Following identification of insufficient stability in human plasma as an important liability of the lead compounds, replacement of the spirosuccinimide core with a spirohydantoin or spiropyrrolidinone structure has yielded a compound that is fully stable in human plasma and with good potency in a human whole blood assay (IC50 = 69 nM) but shows a much lower oral bioavailability (6-9% in rodents) than the earlier compounds. Successive optimization aimed at restoring an acceptable oral bioavailability has yielded compound (S)-17a, which exhibits both stability in human plasma and a good oral bioavailability in rat (37%) and mouse (39%). This compound is also active in a mouse model of ovalbumin-induced lung inflammation following oral dosing at 30 mg/kg.

A constrained reconstruction technique of hyperelasticity parameters for breast cancer assessment.

In breast elastography, breast tissue usually undergoes large compression resulting in significant geometric and structural changes. This implies that breast elastography is associated with tissue nonlinear behavior. In this study, an elastography technique is presented and an inverse problem formulation is proposed to reconstruct parameters characterizing tissue hyperelasticity. Such parameters can potentially be used for tumor classification. This technique can also have other important clinical applications such as measuring normal tissue hyperelastic parameters in vivo. Such parameters are essential in planning and conducting computer-aided interventional procedures. The proposed parameter reconstruction technique uses a constrained iterative inversion; it can be viewed as an inverse problem. To solve this problem, we used a nonlinear finite element model corresponding to its forward problem. In this research, we applied Veronda-Westmann, Yeoh and polynomial models to model tissue hyperelasticity. To validate the proposed technique, we conducted studies involving numerical and tissue-mimicking phantoms. The numerical phantom consisted of a hemisphere connected to a cylinder, while we constructed the tissue-mimicking phantom from polyvinyl alcohol with freeze-thaw cycles that exhibits nonlinear mechanical behavior. Both phantoms consisted of three types of soft tissues which mimic adipose, fibroglandular tissue and a tumor. The results of the simulations and experiments show feasibility of accurate reconstruction of tumor tissue hyperelastic parameters using the proposed method. In the numerical phantom, all hyperelastic parameters corresponding to the three models were reconstructed with less than 2% error. With the tissue-mimicking phantom, we were able to reconstruct the ratio of the hyperelastic parameters reasonably accurately. Compared to the uniaxial test results, the average error of the ratios of the parameters reconstructed for inclusion to the middle and external layers were 13% and 9.6%, respectively. Given that the parameter ratios of the abnormal tissues to the normal ones range from three times to more than ten times, this accuracy is sufficient for tumor classification.

Virtual fragment screening for novel inhibitors of 6-phosphogluconate dehydrogenase.

The enzyme 6-phosphogluconate dehydrogenase is a potential drug target for the parasitic protozoan Trypanosoma brucei, the causative organism of human African trypanosomiasis. This enzyme has a polar active site to accommodate the phosphate, hydroxyl and carboxylate groups of the substrate, 6-phosphogluconate. A virtual fragment screen was undertaken of the enzyme to discover starting points for the development of inhibitors which are likely to have appropriate physicochemical properties for an orally bioavailable compound. A virtual screening library was developed, consisting of compounds with functional groups that could mimic the phosphate group of the substrate, but which have a higher pKa. Following docking, hits were clustered and appropriate compounds purchased and assayed against the enzyme. Three fragments were identified that had IC50 values in the low micromolar range and good ligand efficiencies. Based on these initial hits, analogues were procured and further active compounds were identified. Some of the fragments identified represent potential starting points for a medicinal chemistry programme to develop potent drug-like inhibitors of the enzyme.

Cellular plasticity of inflammatory myeloid cells in the peritoneal foreign body response.

Implantation of sterile foreign objects in the peritoneal cavity of an animal initiates an inflammatory response and results in encapsulation of the objects by bone marrow-derived cells. Over time, a multilayered tissue capsule develops with abundant myofibroblasts embedded in extracellular matrix. The present study used the transgenic MacGreen mouse to characterize the time-dependent accumulation of monocyte subsets and neutrophilic granulocytes in the inflammatory infiltrate and within the tissue capsule by their differential expression of the csf1r-EGFP transgene, F4/80, and Ly6C. As the tissue capsule developed, enhanced green fluorescent protein-positive cells changed from rounded to spindle-shaped morphology and began to co-express the myofibroblast marker alpha-smooth muscle actin. Expression increased with time: at day 14, 11.13 +/- 0.67% of tissue capsule cells co-expressed these markers, compared with 50.77 +/- 12.85% of cells at day 28. The importance of monocyte/macrophages in tissue capsule development was confirmed by clodronate-encapsulated liposome removal, which resulted in almost complete abrogation of capsule development. These results confirm the importance of monocyte/macrophages in the tissue response to sterile foreign objects implanted in the peritoneal cavity. In addition, the in vivo plasticity of peritoneal macrophages and their ability to transdifferentiate from a myeloid to mesenchymal phenotype is demonstrated.

The peritoneal cavity as a bioreactor for tissue engineering visceral organs: bladder, uterus and vas deferens.

Our objective was to produce avascular, myofibroblast-rich tissue capsules for use as autologous grafts for hollow, smooth muscle-walled visceral organs-bladder, uterus and vas deferens. To produce tissue for grafting, templates of the appropriate shape were implanted in the peritoneal cavities of rats or rabbits. After 2-3 weeks, the templates were removed, the encapsulating myofibroblast-rich tissue harvested and grafted to replace resected segments of bladder, vas deferens or uterus of the same animals in which the tissue was grown. Bladder grafts showed 100% patency after 14 months and had developed a morphology similar to normal bladder. Tubes of myofibroblast tissue grafted unilaterally into resected rabbit vasa deferentia developed a morphology resembling native tissue, with sperm in the ejaculate indicative of normal function. At 12 weeks after grafting, uterine graft tissue had increased in thickness and developed the morphology of normal uterus, with endometrium overlying several layers of smooth muscle cells (myometrium-like) which were interspersed with collagen fibrils; grafted uterine horns supported embryos to the late stages of gestation. This study shows that myofibroblast tissue produced in the peritoneal cavity is sufficiently plastic to permit differentiation of cells into bladder, vas deferens or uterine smooth muscle. As a method for producing autologous graft material for repair/replacement of these organs, this approach has many benefits over conventional and current tissue-engineering strategies.

Engineering tissue tubes using novel multilayered scaffolds in the rat peritoneal cavity.

Our aim was to develop novel scaffolds to engineer tissue tubes of smooth muscle-like cells for autologous grafting. Small diameter tubular poly(lactic acid) scaffolds with randomly distributed, interconnected pores up to 100 mum were produced using a thermally induced phase separation method. The scaffolds were surface modified using various biomolecules via a layer-by-layer deposition technique, and implanted in the peritoneal cavities of rats. Histological analysis of scaffolds 3 weeks after implantation showed fully-developed tissue capsules on their outer surfaces, with macrophage-like cells present throughout the internal spaces. Surfaces coated in Matrigel supported the strongest cellular response whereas multilayer coatings with elastin, collagen I, collagen III, or chitosan outermost showed the lowest levels of cellular interaction. Although differences in capsule thickness and the presence or absence of cellularized layers on the inside and outside surfaces of the scaffolds were observed, none of these biomolecule coatings was able to overcome the foreign body response within the peritoneal cavity, even in the presence of a nonadsorptive HA undercoat.

Development of a composite material phantom mimicking the magnetic resonance parameters of the neonatal brain at 3.0 Tesla.

OBJECTIVE: Development of a composite material phantom, comprised of polyvinyl alcohol cryogel (PVA-C) and an agarose additive, to effectively mimic the magnetic resonance relaxation times (T1 and T2) of neonatal white matter (WM) and gray matter (GM) at 3.0 T. MATERIALS AND METHODS: Samples of PVA-C with and without agarose were prepared with 1 cycle of freezing/thawing. Measurements of T1 and T2, at 3.0 T, were performed on the samples at temperatures ranging from 20 degrees C to 40 degrees C. RESULTS: A sample temperature of 40 degrees C was required to achieve a T1 value sufficiently long to represent neonatal WM. At this temperature, neonatal WM relaxation times required 3% PVA-C with 0.3% agarose, whereas gray matter relaxation times required 8% PVA-C with 1.4% agarose. CONCLUSIONS: By adjusting the sample temperature, polyvinyl alcohol concentration, and agarose concentration, the relaxation times of neonatal brain tissues can be obtained using this composite material.

Abduction and external rotation in shoulder impingement: an open MR study on healthy volunteers initial experience.

PURPOSE: To prospectively evaluate rotator cuff contact with the glenoid in healthy volunteers placed in the unloaded and loaded abduction and external rotation (ABER) positions in an open magnetic resonance (MR) imager. MATERIALS AND METHODS: The study was institutional review board approved and HIPAA compliant, and informed consent was received. Eight male volunteers with no history of shoulder pain or pathology were imaged in a 0.5-T open MR imager. Volunteers were imaged in an unloaded ABER position with the arm at 90 degrees abduction and in a loaded ABER position, with a 1-kg load that produced an average external rotation of 111 degrees+/-6 (standard deviation). Two radiologists graded rotator cuff contact on a three-point scale. Three-dimensional anatomic models generated from the MR images were used to measure distances. Minimum distances were computed between the tendon insertion sites and the glenoid, acromion, and coracoid for the loaded ABER position. Minimum distances were compared by using a paired Student t test. RESULTS: In the unloaded ABER position, contact was seen between the infraspinatus and supraspinatus tendons and the glenoid in all eight volunteers. In the loaded ABER position, contact was also observed between the infraspinatus and supraspinatus and the posterior and posterosuperior glenoid, respectively. Deformation of the infraspinatus on the glenoid was seen in four volunteers, whereas supraspinatus deformation was only seen in one volunteer. The minimum distance between the supraspinatus insertion and acromion in the loaded ABER position decreased significantly (P<.01). Supraspinatus tendon to glenoid and infraspinatus tendon to glenoid minimum distances also decreased significantly (P<.01). CONCLUSION: The unloaded and loaded ABER positions resulted in contact of the supraspinatus and infraspinatus with the glenoid in all volunteers. Distances between the rotator cuff insertion sites and the glenoid decreased in the loaded ABER position.

Vascular grafts and the endothelium.

This article discusses the importance of the endothelium for successful vascular grafts derived from both native arteries and synthetic materials. It also discusses the fundamental strategies to endothelialize synthetic grafts in animal experiments and in the clinic, as well as the use of endothelial progenitor cells (EPCs), bone marrow-derived cells, and mesothelium as endothelial substitutes.

CbrA is a stationary-phase regulator of cell surface physiology and legume symbiosis in Sinorhizobium meliloti.

Sinorhizobium meliloti produces an exopolysaccharide called succinoglycan that plays a critical role in promoting symbiosis with its host legume, alfalfa (Medicago sativa). We performed a transposon mutagenesis and screened for mutants with altered succinoglycan production and a defect in symbiosis. In this way, we identified a putative two-component histidine kinase associated with a PAS sensory domain, now designated CbrA (calcofluor-bright regulator A). The cbrA::Tn5 mutation causes overproduction of succinoglycan and results in increased accumulation of low-molecular-weight forms of this exopolysaccharide. Our results suggest the cbrA::Tn5 allele leads to this succinoglycan phenotype through increased expression of exo genes required for succinoglycan biosynthesis and modification. Interestingly, CbrA-dependent regulation of exo and exs genes is observed almost exclusively during stationary-phase growth. The cbrA::Tn5 mutant also has an apparent cell envelope defect, based on increased sensitivity to a number of toxic compounds, including the bile salt deoxycholate and the hydrophobic dye crystal violet. Growth of the cbrA mutant is also slowed under oxidative-stress conditions. The CbrA-regulated genes exsA and exsE encode putative inner membrane ABC transporters with a high degree of similarity to lipid exporters. ExsA is homologous to the Escherichia coli MsbA protein, which is required for lipopolysaccharide transport, while ExsE is a member of the eukaryotic family of ABCD/hALD peroxisomal membrane proteins involved in transport of very long-chain fatty acids, which are a unique component of the lipopolysaccharides of alphaproteobacteria. Thus, CbrA could play a role in regulating the lipopolysaccharide or lipoprotein components of the cell envelope.

Sinorhizobium meliloti bluB is necessary for production of 5,6-dimethylbenzimidazole, the lower ligand of B12.

An insight into a previously unknown step in B(12) biosynthesis was unexpectedly obtained through our analysis of a mutant of the symbiotic nitrogen fixing bacterium Sinorhizobium meliloti. This mutant was identified based on its unusually bright fluorescence on plates containing the succinoglycan binding dye calcofluor. The mutant contains a Tn5 insertion in a gene that has not been characterized previously in S. meliloti. The closest known homolog is the bluB gene of Rhodobacter capsulatus, which is implicated in the biosynthesis of B(12) (cobalamin). The S. meliloti bluB mutant is unable to grow in minimal media and fails to establish a symbiosis with alfalfa, and these defects can be rescued by the addition of vitamin B(12) (cyanocobalamin) or the lower ligand of cobalamin, 5,6-dimethylbenzimidazole (DMB). Biochemical analysis demonstrated that the bluB mutant does not produce cobalamin unless DMB is supplied. Sequence comparison suggests that BluB is a member of the NADH/flavin mononucleotide (FMN)-dependent nitroreductase family, and we propose that it is involved in the conversion of FMN to DMB.