Human responses to propionic acid. I. Quantification of within- and between-participant variation in perception by normosmics and anosmics

The objective of this study was to fully characterize normosmic perception of stimuli expected to cause widely varying degrees of olfactory and nasal trigeminal stimulation and to directly evaluate the possible role of olfactory nerve stimulation in nasal irritation sensitivity. During each of four identical test sessions, four anosmic and 31 normosmic participants were presented with a range of concentrations extending from peri-threshold for normosmics to supra- threshold for anosmics. For each session, odor (O) and nasal irritation (NI) sensitivities were summarized in terms of the concentrations required to produce four sensation levels ('iso-response' concentrations). Within-participant variation in these iso-response concentrations was < 10-fold for 95% of normosmics, for both O and NI. For O but not NI, these apparent fluctuations in sensitivity were largely accounted for by the uncertainty surrounding the iso-response concentrations calculated for each session. Anosmics exhibited minimal within- and between-participant variation in NI and required, for all but the highest perceptual level, a higher concentration than almost all normosmics. Between-participant variation, expressed in terms of 90% confidence interval widths, was approximately 0.5 log units for both O and NI for the highest perceptual level, but increased to approximately 0.8 and 1.8 log units, respectively, for the lowest (peri- threshold) level. Our findings suggest that: (i) most apparent variation over time in O sensitivity is actually a reflection of the uncertainty surrounding estimates of sensitivity obtained for each session; (ii) within- and between-participant variation in O sensitivity is far less than is commonly reported; and (iii) low to moderate levels of NI in normosmics are the result of relatively weak trigeminal stimulation combined with much greater olfactory activation.      

Human breathing pattern responses to loading with increased background impedance

We determined the influence of the background level of mechanical impedance on the respiratory responses to very small mechanical loads, at or below the threshold for conscious perception. We used a pseudorandom load application technique to estimate the immediate pattern responses from the zeroth lag of the cross correlation between the load application sequence and the respiratory pattern components of tidal volume (VT), inspiratory and expiratory time (TI and TE), and the instantaneous respiratory frequency (f), minute ventilation (VI), and mean inspiratory flow (VT/TI). Elevation of the background resistance served to reduce the TI and TE responses to small perturbations in resistance from those in the control background state, which resulted in generally smaller perturbations of f, VI, and VT/TI. Elevation of the background elastance, however, served to initiate a TI reduction not seen in the control state but did not appreciably affect the rest of the pattern responses to the load perturbations. Thus the neural reflexes involved in breath-by-breath pattern regulation are modulated by the background level of the respiratory impedance, as well as by the type and size of the load perturbation.     

Fluoroquinolones and propionic acid derivatives induce inflammatory responses in vitro

Fluoroquinolones and propionic acid derivatives are widely used antibacterials and non-steroidal anti-inflammatory drugs, respectively, which have been reported to frequently trigger drug hypersensitivity reactions. Such reactions are induced by inflammatory mediators such as cytokines and chemokines. The present study investigated whether levofloxacin, a fluoroquinolone, and loxoprofen, a propionic acid derivative, have the potential to induce immune-related gene expression in dendritic cell-like cell lines such as HL-60, K562, and THP-1, and immortalized keratinocytes such as HaCaT. The expression of IL-8, MCP-1, and TNFα messenger RNA (mRNA) was found to increase following treatment with levofloxacin or loxoprofen in HL-60 cells. In addition, these drugs increased the mRNA content of annexin A1, a factor related to keratinocyte necroptosis in patients with severe cutaneous adverse reactions. Inhibition studies using specific inhibitors of mitogen-activated protein (MAP) kinases and NF-κB suggest that the extracellular signal-regulated kinase (ERK) pathway is the pathway principally involved in the induction of cytokines and annexin A1 by levofloxacin, whereas the involvement of MAP kinases and NF-κB in the loxoprofen-induced gene expression of these factors may be limited. Fluoroquinolones and propionic acid derivatives that are structurally related to levofloxacin and loxoprofen, respectively, were also found to induce immune-related gene expression in HL-60 cells. Collectively, these results suggest that fluoroquinolones and propionic acid derivatives have the potential to induce the expression of immune-related factors and that an in vitro cell-based assay system to detect the immune-stimulating potential of systemic drugs might be useful for assessing the risk of drug hypersensitivity reactions.     

Antimutagenicity of propionic acid bacteria.

The antimutagenic effect of dialysed cell extracts of 4 strains of propionic acid bacteria was examined against the mutagenicity of sodium azide in the TA1535 tester strain of Salmonella typhimurium using the Ames test. It was noted that dialysates of 2 strains of Propionibacterium shermanii, P. pentosaceum and P. acnes, significantly reduced sodium azide-induced revertants. The dialysate of propionic acid cocci did not show an antimutagenic effect. The inhibitory activity was enhanced if the mutagen and extract were coincubated for 20 min prior to performing the mutagenicity assay. Antimutagenicity of dialysates from P. shermanii VKM-103 against MNNG and 9-aminoacridine was shown in S. typhimurium strains TA1535 and TA97. The antimutagenic activity was found in the protein fraction of the cell extract of P. shermanii. The proteins of the dialysate of P. shermanii were separated using a Toyopearl gel column into 3 main peaks according to their molecular weights. The antimutagenic activity towards sodium azide was found in the second and the third peaks. We suggest that dialysates of the cells of propionic acid bacteria contain several kinds of antimutagenic substances with different molecular weights.     

Human evoked responses to potentially noxious tactile stimulation, II.

Signal summation and minimally adapting tactile stimulation techniques permit the resolution of a critical time domain of the somatosensory evoked response, recorded from human scalp, which is sensitive to the stimulus variables represented by the sensations of tapping and pinprick respectively. The variation of this time-domain suggests the influence of myelinated mechanical nociceptors in the Group III range.     

Boophilus microplus: cellular responses to larval attachment and their relationship to host resistance.

The histology of early feeding lesions of the cattle tick B. microplus has been studied using 32P labelled larvae to standardize the duration of attachment. Critical studies were made on 3-h lesions in six separate experiments on different groups of British breed animals. Each group consisted of three animals--one previously unexposed to ticks, one of high resistance and one of low resistance. The degree of mast cell disruption, eosinophil concentration and degranulation, and the extent of epidermal vesiculation were all significantly greater at the site of attachment on highly resistant hosts. In previously unexposed animals there was no mobilization of eosinophils nor mast cell breakdown and no epidermal vesiculation. Possible immune mechanisms producing mast cell disruption and the infiltration and concentration of eosinophils are suggested, and the effect of eosinophil degranulation on larval attachment and feeding is discussed.     

Comparison of identified mitral and tufted cells in freely breathing rats: II. Odor-evoked responses

Mitral and tufted cells are the 2 types of output neurons of the main olfactory bulb. They are located in distinct layers, have distinct projection patterns of their dendrites and axons, and likely have distinct relationships with the intrabulbar inhibitory circuits. They could thus be functionally distinct and process different aspects of olfactory information. To examine this possibility, we compared the odor-evoked responses of identified single units recorded in the mitral cell layer (MCL units), in the core of the external plexiform layer (not at the glomerular border tufted cells), or at the glomerular border of this layer (GB tufted cells) of the entire olfactory bulb. Differences between mitral and tufted cells were observed only when subtle aspects of the responses were explored, such as the firing rate per respiratory cycle or the distribution of firing activity along the respiratory cycle. By contrast, more clear differences were found when the 2 subtypes of tufted cells were examined separately. GB units were significantly more responsive, had significantly higher firing activity, and showed greater activity at the transition between inspiration and expiration. The projection-type tufted cells situated closer to the entrance of the olfactory bulb may thus form a distinct physiological class of output neurons and differ from mitral cells and other tufted cells in the manner of processing olfactory information.     

Quantification of free ligand conformational preferences by NMR and their relationship to the bioactive conformation

Accurate unbound solution 3D-structures of ligands provide unique opportunities for medicinal chemistry and, in particular, a context to understand binding thermodynamics and kinetics. Previous methods of deriving these 3D-structures have had neither the accuracy nor resolution needed for drug design and have not yet realized their potential. Here, we describe and apply a NMR methodology to the aminoglycoside streptomycin that can accurately quantify accessible 3D-space and rank the occupancy of observed conformers to a resolution that enables medicinal chemistry understanding and design. Importantly, it is based upon conventional small molecule NMR techniques and can be performed in physiologically-relevant solvents. The methodology uses multiple datasets, an order of magnitude more experimental data than previous NMR approaches and a dynamic model during refinement, is independent of computational chemistry and avoids the problem of virtual conformations. The refined set of solution 3D-shapes for streptomycin can be grouped into two major families, of which the most populated is almost identical to the 30S ribosomal subunit bioactive shape. We therefore propose that accurate unbound ligand solution conformations may, in some cases, provide a subsidiary route to bioactive shape without crystallography. This experimental technique opens up new opportunities for drug design and more so when complemented with protein co-crystal structures, SAR data and pharmacophore modeling.     

Human immune responses to hapten-conjugated cells. II. The roles of autologous and allogeneic histocompatibility determinants in proliferative responses in vitro.

Proliferative responses of human lymphocytes primed in vitro to autologous TNP-cells were found to be associated with autologous D-region determinants irrespective of HLA-B locus antigens. Family studies of secondary TNP-conjugate proliferative responses demonstrated a gene dosage effect in this phenomenon. Moreover, co-culture with allogeneic cells did not affect the net TNP-conjugate proliferative responses of primed responder cells, suggesting that HLA-D region preference was due to a requirement for representation of TNP-molecules in association or combination with autologous MHC structures. Alloantigens were found to influence the sensitization of lymphocytes to autologous hapten-conjugated cells. Co-culture of allogeneic and TNP-modified autologous stimulator cells in primary cultures enhanced the secondary TNP proliferative response. Sensitization of human lymphocytes to allogeneic cells alone did not prime responses to autologous modified cells. However, priming lymphocytes to modified autologous cells potentiated responses to allogeneic cells. The data suggest a complex relationship between responses to alloantigens and modified autologous cells.     

Chromium(III) complexes and their relationship to the glucose tolerance factor. Part II. Structure and biological activity of amino acid complexes

A number of octahedral chromium complexes with amino acids are ligands have been prepared and their structures assigned on the basis of their chromatographic and spectral properties. These include complexes with the general structure Cr(AA)₂(H₂O)₂ where the amino acids glycine, glutamic acid and glutamine act as bidentate ligands. The analogous compound with cysteine as ligand is stable at low pH, but at high pH a terdentate cysteine complex, Cr(cysteine)₂^?, is formed. These complexes, as well as a solution of monodentate glycine aquo complexes, and Cr-nicotinic acid-glycine and Cr-nicotinic acid-cysteine complexes of undetermined structure, have been assayed for glucose tolerance factor activity using a yeast assay. Only Cr(glutamine)₂- (H₂O)₂⁺, Cr-nicotinic acid-glycine and the mixture of complexes Cr(glycine)_n(H₂O)_6-n⁺³ showed significant activity. It is proposed that a trans arrangement of the non-coordinated nitrogen atoms in the ligands of these complexes can mimic the structural features of the glucose tolerance factor which are essential for biological activity.     

Clostridium sporogenes isolates and their relationship to C. botulinum based on deoxyribonucleic acid reassociation.

Sixty-two isolates of Clostridium sporogenes from canned foods were examined for cultural properties, heat resistance and DNA-DNA homology to Clostridium botulinum type A190. Sporulation was observed in most of 21 umbonate and rhizoidal colony-forming strains (colony-type I strains), but not in most of the 41 strains with convex and circular or crenate colonies with a mat to semi-glossy surface (colony-type II strains). More than half of the latter strains showed much higher heat resistance than the rhizoidal colony-forming strains. The DNA isolated from colony-type II strains was 81% or more homologous to C. botulinum A190 DNA, forming duplexes which had thermostabilities similar to homologous duplexes of strain A190 DNA. Colony-type I strains differed from C. botulinum by 30 to 40% DNA homology and the DNA duplexes formed between these strains and strain A190 showed deltaT m(e) values of 7-0 degrees C when compared with the T m(e) of homologous DNA duplexes of strain A190.     

Human eosinophil cytotoxicity-enhancing factor. II. Multiple forms synthesized by U937 cells and their relationship to thioredoxin/adult T cell leukemia-derived factor

Recently, our laboratory reported the purification and partial amino acid sequence of a 10-kDa eosinophil cytotoxicity-enhancing factor (ECEF) polypeptide from the U937 cell source. This cytokine enhanced human eosinophil antibody-dependent cytotoxic function by greater than 200% and was half-maximally effective at a concentration of approximately 1 ng/ml. In this study, we describe the conditions required for ECEF synthesis and the use of rabbit antibody raised to 10-kDa ECEF to investigate the existence of related polypeptide species. Unstimulated U937 cells released an immunoreactive 14-kDA species. Cells stimulated with 7.5 micrograms/ml of LPS also released a 13-kDa species. Cells stimulated with 400 ng/ml of PMA also synthesized a 10-kDa species (equivalent in size to the form we had purified). This 10-kDa species remained primarily cell associated, but detectable amounts were released into the supernatant by 48 h of culture. In washed cell pellets, the location of the 10-kDa species was found to be in the plasma membrane, externally oriented, as determined by FACS analysis, iodination with the membrane impermeable reagent 125I-sulfosuccinimidyl-3-(4-hydroxyphenyl) propionate, and by its removal with brief trypsin treatment. Partial amino acid sequence data suggested that the 14-, 13-, and 10-kDa species all share the same N-terminal. The 14- and 10-kDa ECEF species were recovered by electroelution from SDS-PAGE gels and tested for activity in the assay of eosinophil cytotoxic function. Because of the amino acid sequence similarities between the ECEF species and thioredoxin (TRX), rTRX (synthesized in Escherichia coli and purified) was also tested for activity. The 14-kDa ECEF and rTRX induced a slight, but consistent and statistically significant enhancement of eosinophil cytotoxic function. By comparison, lower doses of the 10-kDa ECEF induced a major increase in cytotoxic function. Thus the forms of ECEF differ in size, conditions required for synthesis, trafficking by the U937 cell after synthesis, and biologic activity. It is likely that these considerations bear on the involvement of ECEF in the pathophysiology of eosinophilia in vivo.     

Structure-activity relationship of triaryl propionic acid analogues on the human EP3 prostanoid receptor

Potent and selective ligands for the human EP3 prostanoid receptor are described. Triaryl compounds bearing an ortho-substituted propionic acid moiety were identified as potent EP3 antagonists based on the SAR described herein. The binding affinities of key compound on all eight human prostanoid receptors is reported.