Heat stress associated with aerosol PPE and its impact.

BACKGROUND: Aerosol personal protective equipment (PPE) is subjectively reported to negatively impact healthcare workers' performance and well-being, but this has not been assessed objectively. AIMS: This randomized controlled crossover study aimed to quantify the heat stress associated with aerosol PPE and to investigate its impact upon mood, cognitive and motor function, and task performance. METHODS: Sixteen healthy, young, lean participants (eight males) undertook an exercise protocol, which simulated the metabolic expenditure of hospital work: once wearing aerosol PPE (PPE visit) and once wearing standard surgical attire (control visit). Participants walked on a treadmill for 2 h followed by 30-min rest. Core temperature, heart rate, urine specific gravity, weight, grip strength, mood (Bond-Lader scale) and task performance (Intubation of a Manikin) were recorded. Values are between-visit mean (standard deviation) differences. RESULTS: On the PPE visit core temperature (+0.2 (0.3)°C; P < 0.01), heart rate (+12 (13) bpm; P < 0.001), urine specific gravity (+0.003 (0.005); P < 0.05) and intubation task time (+50 (81) s; P < 0.01) were greater than on the control visit; and alertness (-14 (21) mm; P < 0.001), contentment (-14 (15) mm; P < 0.001) and grip strength (-4 (4) N; P < 0.01) were less. CONCLUSIONS: This study demonstrates that wearing aerosol PPE in a simulated hospital environment results in heat exhaustion and has a negative impact upon mood, motor function, and task performance. Whilst wearing PPE is important to prevent disease transmission, strategies should be developed to limit its impact upon healthcare workers' performance and well-being.

Coherency image analysis to quantify collagen architecture: implications in scar assessment.

An important histological difference between normal, uninjured dermis and scar tissue such as that found in keloid scars is the pattern (morphological architecture) in which the collagen is deposited and arranged. In the uninjured dermis, collagen bundle architecture appears randomly organized (or in a basket weave formation), whereas in pathological conditions such as keloid scar tissue, collagen bundles are often found in whorls or in a hypotrophic scar collagen is more densely packed in a parallel configuration. In the case of skin, a scar disables the dermis, leaving it weaker, stiff and with a loss of optimal functionality. The absence of objective and quantifiable assessments of collagen orientation is a major bottleneck in monitoring progression of scar therapeutics. In this article, a novel quantitative approach for analyzing collagen orientation is reported. The methodology is demonstrated using collagen produced by cells in a model scar environment and examines collagen remodeling post-TGFß stimulation in vitro. The method is shown to be reliable and effective in identifying significant coherency differences in the collagen deposited by human keloid scar cells. The technique is also compared for analysing collagen architecture in rat sections of normal, scarred skin and tendon tissue. Results demonstrate that the proposed computational method provides a fast and robust way of analyzing collagen orientation in a manner surpassing existing methods. This study establishes this methodology as a preliminary means of monitoring in vitro and in tissue treatment modalities which are expected to alter collagen morphology.

Maghemite (hematite) core (shell) nanorods via thermolysis of a molecular solid of Fe-complex.

An Fe-metal complex with 2'-hydroxy chalcone (2'-HC) ligands [Fe(III) (2'-hydroxy chalcone)(3)] is synthesized by a chemical route and is subjected to different thermal treatments. Upon thermolysis in air at 450 °C for 3 h the complex yields maghemite (γ-Fe(2)O(3)) nanorods with a thin hematite (α-Fe(2)O(3)) shell. X-Ray diffraction (XRD), Mössbauer spectroscopy, diffuse reflectance spectroscopy (UV-DRS), high resolution transmission electron microscopy (HR-TEM), field emission scanning electron microscopy (FE-SEM) and vibrating sample magnetometry (VSM) are used to characterize the samples. The stability of the ligand and the Fe-complex is further examined by using thermogravimmetric/differential thermal analysis (TGA/DTA). We suggest a residual ligand controlled mechanism for the formation of an anisotropic nanostructure in a crumbling molecular solid undergoing ligand decomposition. Since the band gap of iron oxide is in the visible range, we explored the use of our core shell nano-rod sample for photocatalytic activity for H(2) generation by H(2)S splitting under solar light. We observed high photocatalytic activity for hydrogen generation (75 ml h(-1)).

A metabotropic glutamate receptor variant functions as a taste receptor.

Sensory transduction for many taste stimuli such as sugars, some bitter compounds and amino acids is thought to be mediated via G protein-coupled receptors (GPCRs), although no such receptors that respond to taste stimuli are yet identified. Monosodium L-glutamate (L-MSG), a natural component of many foods, is an important gustatory stimulus believed to signal dietary protein. We describe a GPCR cloned from rat taste buds and functionally expressed in CHO cells. The receptor couples negatively to a cAMP cascade and shows an unusual concentration-response relationship. The similarity of its properties to MSG taste suggests that this receptor is a taste receptor for glutamate.

An optimized method for in situ hybridization with signal amplification that allows the detection of rare mRNAs.

In situ hybridization (ISH) using nonradioactive probes enables mRNAs to be detected with improved cell resolution but compromised sensitivity compared to ISH with radiolabeled probes. To detect rare mRNAs, we optimized several parameters for ISH using digoxygenin (DIG)-labeled probes, and adapted tyramide signal amplification (TSA) in combination with alkaline phosphatase (AP)-based visualization. This method, which we term TSA-AP, achieves the high sensitivity normally associated with radioactive probes but with the cell resolution of chromogenic ISH. Unlike published protocols, long RNA probes (up to 2.61 kb) readily permeated cryosections and yielded stronger hybridization signals than hydrolyzed probes of equivalent complexity. RNase digestion after hybridization was unnecessary and led to a substantial loss of signal intensity without significantly reducing nonspecific background. Probe concentration was also a key parameter for improving signal-to-noise ratio in ISH. Using these optimized methods on rat taste tissue, we detected mRNA for mGluR4, a receptor, and transducin, a G-protein, both of which are expressed at very low abundance and are believed to be involved in chemosensory transduction. Because the effect of the tested parameters was similar for ISH on sections of brain and tongue, we believe that these methodological improvements for detecting rare mRNAs may be broadly applicable to other tissues. (J Histochem Cytochem 47:431-445, 1999)

Molecular and physiological evidence for glutamate (umami) taste transduction via a G protein-coupled receptor.

Recent molecular analyses have demonstrated that a metabotropic glutamate receptor, mGluR4, is expressed in taste buds from rat circumvallate and foliate papillae. Behavioral studies demonstrated that L(+)-2-amino-4-phosphonobutyric acid (L-AP4), an agonist for mGluR4 and related receptors, mimics the taste of monosodium glutamate (MSG) in rats. mGluR4 is known to signal through inhibition of the cyclic adenosine-5',3'-monophosphate (cAMP) cascade. Circumvallate and foliate taste buds exhibit decreases of cAMP levels following stimulation with MSG, and the response is potentiated by 5'-inosine monophosphate, suggesting that it is related to umami taste. Further, experiments on mice with the mGluR4 gene knocked out support the interpretation that mGluR4 is a key component in glutamate taste. Glutamate may also stimulate taste buds through an ionotropic receptor pathway. In patch-clamp studies, glutamate evokes two types of currents, similar to those elicited by N-methyl-D-aspartate (NMDA) and L-AP4. We speculate upon the significance of two glutamate receptor pathways in taste buds.

Responses to glutamate in rat taste cells.

We studied taste transduction in sensory receptor cells. Specifically, we examined the actions of glutamate, a significant taste stimulus, on the membrane properties of taste cells by applying whole cell patch-clamp techniques to cells in rat taste buds isolated from foliate and vallate papillae. In 55 of 91 taste cells, bath-applied glutamate, at concentrations that elicit taste responses in the intact animal (10-20 mM), produced one of two different responses when the cell membrane was held near its presumed resting potential, -85 mV. "Sustained" glutamate responses were observed in the majority of taste cells (51 of 55) and consisted of an outward current (reduction of the maintained inward current). Sustained glutamate responses were voltage dependent, were decreased by membrane depolarization, and were accompanied by a reduction in membrane conductance. An analysis of the reversal potential of sustained responses in different ionic conditions and the effect of ion substitutions suggested that the currents were carried by cations. The data suggest that sustained responses are mediated by the closure of nonselective cation channels. Other taste cells (4 of 55) responded to glutamate with a transient inward current--so-called "transient" responses. Transient glutamate responses were voltage dependent and Na+ dependent, and appeared to be generated by nonspecific cation channels activated by glutamate. L(+)-2-amino-4-phosphonobutyric acid (L-AP4), a specific agonist of a metabotropic glutamate receptor (mGluR4) recently identified in rat taste cells and believed to be involved in taste transduction, mimicked the sustained glutamate responses. These findings indicate that glutamate, at concentrations at or slightly above threshold for taste in rats, produces two different membrane currents. The properties of these two responses suggest that there may be two different sets of nonspecific cation channels in taste cells, one closed by glutamate (sustained response) and the other opened (transient response). Our findings on the effect of L-AP4 suggest that the sustained response is the membrane mechanism mediating, at least in part, taste transduction for glutamate.

The taste of monosodium glutamate: membrane receptors in taste buds.

Receptor proteins for photoreception have been studied for several decades. More recently, putative receptors for olfaction have been isolated and characterized. In contrast, no receptors for taste have been identified yet by molecular cloning. This report describes experiments aimed at identifying a receptor responsible for the taste of monosodium glutamate (MSG). Using reverse transcriptase (RT)-PCR, we found that several ionotropic glutamate receptors are present in rat lingual tissues. However, these receptors also could be detected in lingual tissue devoid of taste buds. On the other hand, RT-PCR and RNase protection assays indicated that a G-protein-coupled metabotropic glutamate receptor, mGluR4, also is expressed in lingual tissues and is limited only to taste buds. In situ hybridization demonstrated that mGluR4 is detectable in 40-70% of vallate and foliate taste buds but not in surrounding nonsensory epithelium, confirming the localization of this metabotropic receptor to gustatory cells. Expression of mGluR4 in taste buds is higher in preweaning rats compared with adult rats. This may correspond to the known higher sensitivity to the taste of MSG in juvenile rodents. Finally, behavioral studies have indicated that MSG and L-2-amino-4-phosphonobutyrate (L-AP4), a ligand for mGluR4, elicit similar tastes in rats. We conclude that mGluR4 may be a chemosensory receptor responsible, in part, for the taste of MSG.

Knowledge representation using fuzzy deduction graphs.

A new knowledge representation model, known as fuzzy deduction graph (FDG), is introduced in this paper. An FDG can represent a knowledge base containing the fuzzy propositions and fuzzy rules. In an FDG, a systematic method of finding the fuzzy reasoning path (FRP) is given which is based on Dijkstra's shortest path framework. The FRP gives a relationship between the antecedent (source) proposition and consequent (goal) proposition, such that the consequent proposition is reached with the greatest fuzzy value. The process of finding the FRP is illustrated with examples.

mRNA for cardiac calcium channel is expressed during development of skeletal muscle.

During the early development of skeletal muscle, cardiac isotypes of several contractile proteins are known to be transiently expressed. We report here that skeletal muscle developing in vivo, as well as primary cultures derived from skeletal muscle, express mRNA encoding the cardiac dihydropyridine-sensitive calcium channel. The mRNA is detectable at high concentration at the earliest stage tested in vivo and diminishes rapidly in concentration as myofibers mature. The concentration of the cardiac calcium channel mRNA also diminishes during the in vivo development of skeletal muscle in a genetically paralyzed mouse (mdg), indicating that muscle contractile activity is not necessary for the down-regulation. In contrast, mRNA for the skeletal muscle-specific calcium channel accumulates gradually in developing skeletal muscle. A similar temporal pattern of expression is also seen in primary cultures of skeletal myotubes. These results raise the question of whether the cardiac calcium channel may be functionally important during the early development of skeletal myofibers.

A single nucleotide deletion in the skeletal muscle-specific calcium channel transcript of muscular dysgenesis (mdg) mice.

The skeletal muscle-specific dihydropyridine-sensitive calcium channel is a critical component of excitation-contraction coupling in skeletal muscle. A recessive mutation in mice, muscular dysgenesis (mdg), has previously been described as resulting in defective excitation-contraction coupling. Although the channel-forming subunit (alpha 1) of the skeletal calcium channel is not detectable immunologically, specific mRNA of normal size is present in dysgenic muscle. cDNA for this calcium channel alpha 1 subunit has now been cloned from dysgenic muscle and sequenced in its entirety. A single nucleotide deletion occurs at nucleotide 4010 of the cDNA, resulting in a shift of the translational reading frame. The mutation has been confirmed by direct sequencing of PCR products from homozygous mutant and normal muscle. The mutant polypeptide is predicted to contain the first three repeating domains, five of the normal six transmembrane helices of the last repeating domain, and an altered and truncated C terminus. The mature mRNA encoding the dysgenic alpha 1 subunit appears to be labile. It is possible that premature termination of translation renders the mutant mRNA subject to degradation by nucleases. This work resolves a long-standing controversy on the nature of the primary genetic defect in muscular dysgenesis.

Restoration of normal function in genetically defective myotubes by spontaneous fusion with fibroblasts.

Muscular dysgenesis in mice is a genetic disease of skeletal muscle caused by the recessive mutation mdg. Muscle fibres in affected mice are paralysed because of the failure of excitation-contraction coupling. Unlike normal myotubes in primary culture, dysgenic myotubes do not contract, either spontaneously or in response to electrical stimulation. The deficiency results from mutation of the gene for the skeletal muscle dihydropyridine receptor, an essential sarcolemmal component both of excitation-contraction coupling and of the slow calcium-ion channel. It has recently been shown that the addition of fibroblasts from normal (but not dysgenic) mice to cultures of dysgenic myotubes can restore spontaneous contractions in a small fraction of these myotubes, but the mechanism for this 'rescue' was not determined. In principle, if fibroblast nuclei were able to incorporate into myotubes, such nuclei could then supply the missing muscle-specific gene product. We have now investigated this possibility using nuclear, cytoplasmic and plasmalemmal markers. We report that the rescue to contractile ability in genetically paralysed dysgenic muscle is mediated by the previously unrecognized ability of fibroblasts to fuse spontaneously with developing myotubes.

The muscular dysgenesis mutation in mice leads to arrest of the genetic program for muscle differentiation.

Muscular dysgenesis (mdg) is a mutation in mice which causes the failure of excitation-contraction coupling in skeletal muscle. Although the sarcolemma, the sarcoplasmic reticulum, and the contractile apparatus all maintain nearly normal function, sarcolemmal depolarization fails to cause calcium release from the sarcoplasmic reticulum. Recently, the primary genetic defect in this mutation was shown to be located in the structural gene for the dihydropyridine receptor. We have examined the developmental expression from Fetal Day 15 onward, in normal and mutant muscle, of several unidentified genes as well as genes which are known markers of muscle differentiation. We find that the majority of mRNA sequences are found at similar concentrations in normal and dysgenic muscles at birth. Many differentiation-related genes also are expressed at normal levels early during myogenesis in mutant mice. However, as late fetal development progresses in dysgenic muscle, the mRNA concentrations for these genes fail to undergo the rapid rise which is characteristic of normal muscle. Several additional, unidentified genes, which normally would be down-regulated during development, remain expressed at a high level in dysgenic muscle. Thus, the primary absence of a functional dihydropyridine receptor appears to prevent the changes in gene expression which are necessary for maturation of skeletal muscle.

Specific absence of the alpha 1 subunit of the dihydropyridine receptor in mice with muscular dysgenesis.

Muscular dysgenesis is a lethal mutation in mice that results in a complete absence of skeletal muscle contraction due to the failure of depolarization of the transverse tubular membrane to trigger calcium release from the sarcoplasmic reticulum. In order to determine whether the defect in muscular dysgenesis leads to a specific loss of one of the components of excitation-contraction coupling or to a generalized loss of all components of excitation-contraction coupling, we have analyzed skeletal muscle from control and dysgenic mice for the sarcoplasmic reticulum and transverse tubular proteins which are believe to function in excitation-contraction coupling. We report that the proteins involved in sarcoplasmic reticulum calcium transport, storage, and release [Ca2+ + Mg2+)-ATPase, calsequestrin, and calcium release channel) are present in dysgenic muscle. Also present in dysgenic muscle is the 175/150-kDa glycoprotein subunit (alpha 2) of the dihydropyridine receptor. However, the 170-kDa dihydropyridine binding subunit (alpha 1) of the dihydropyridine receptor is absent in dysgenic muscle. These results suggest that the specific absence of the alpha 1 subunit of the dihydropyridine receptor is responsible for the defects in muscular dysgenesis and that the alpha 1 subunit of the dihydropyridine receptor is essential for excitation-contraction coupling in skeletal muscle.

Comparison of pulsatile subcutaneous gonadotropin-releasing hormone and exogenous gonadotropins in the treatment of men with isolated hypogonadotropic hypogonadism.

Eight men with isolated hypogonadotropic hypogonadism were treated with pulsatile gonadotropin-releasing hormone (GnRH) after maximal testicular growth and function had already been achieved with human chorionic gonadotropin (hCG) and human menopausal gonadotropin (hMG). Only four subjects could normalize plasma testosterone (T) levels (group A). After 18 months of GnRH therapy, testicular size of group A increased by 53% (P less than 0.01) over that previously attained with exogenous gonadotropins. However, despite further testicular growth, two men who were previously azoospermic on hCG/hMG remained so on GnRH. In the other two patients, total sperm count increased minimally. Thus, pulsatile gonadotropin levels achieved with GnRH are more effective in stimulating testicular growth, but not necessarily sperm output, than are stable gonadotropin concentrations obtained with hCG/hMG.