Role of GDP in formyl-peptide-receptor-induced activation of guanine-nucleotide-binding proteins in membranes of HL 60 cells.

Membranes of myeloid differentiated human leukemia (HL 60) cells contain receptors for the chemotactic peptide, fMet-Leu-Phe (fMet, N-formylmethionine), interacting with pertussis-toxin-sensitive guanine-nucleotide-binding proteins (G proteins). Agonist activation of the receptors increases binding of the GTP analog, guanosine 5'-[gamma-thio]triphosphate (GTP[S]), to membrane G proteins, at 30 degrees C only in the presence of exogenous GDP. In contrast, at 0 degrees C fMet-Leu-Phe stimulated binding of GTP[S] to G proteins maximally without addition of GDP. Under conditions resulting in marked degradation of membrane-bound GDP, control binding of GTP[S] measured at 0 degrees C was significantly increased, whereas the extent of agonist-stimulated binding was reduced. Furthermore, there was a rapid spontaneous release of membrane-bound GDP at 30 degrees C, but not at 0 degrees C. The data suggest that in intact membranes of HL 60 cells G proteins are initially in a GDP-liganded form, which state allows the receptor-induced exchange of bound GDP for GTP[S] at low temperature. In contrast, at or near physiological temperature, bound GDP is rapidly released (and degraded), resulting in unligated G proteins to which GTP[S] will bind independently of agonist-activated receptors.     

Increased Respiratory Disease Mortality at a Microwave Popcorn Production Facility with Worker Risk of Bronchiolitis Obliterans

Background

Bronchiolitis obliterans, an irreversible lung disease, was first associated with inhalation of butter flavorings (diacetyl) in workers at a microwave popcorn company. Excess rates of lung-function abnormalities were related to cumulative diacetyl exposure. Because information on potential excess mortality would support development of permissible exposure limits for diacetyl, we investigated respiratory-associated mortality during 2000–2011 among current and former workers at this company who had exposure to flavorings and participated in cross-sectional surveys conducted between 2000–2003.

Methods

We ascertained workers'' vital status through a Social Security Administration search. Causes of death were abstracted from death certificates. Because bronchiolitis obliterans is not coded in the International Classification of Disease 10^th revision (ICD-10), we identified respiratory mortality decedents with ICD-10 codes J40–J44 which encompass bronchitis (J40), simple and mucopurulent chronic bronchitis (J41), unspecified chronic bronchitis (J42), emphysema (J43), and other chronic obstructive pulmonary disease (COPD) (J44). We calculated expected number of deaths and standardized mortality ratios (SMRs) with 95% confidence intervals (CI) to determine if workers exposed to diacetyl experienced greater respiratory mortality than expected.

Results

We identified 15 deaths among 511 workers. Based on U.S. population estimates, 17.39 deaths were expected among these workers (SMR = 0.86; CI:0.48-1.42). Causes of death were available for 14 decedents. Four deaths among production and flavor mixing workers were documented to have a multiple cause of ‘other COPD’ (J44), while 0.98 ‘other COPD’-associated deaths were expected (SMR = 4.10; CI:1.12–10.49). Three of the 4 ‘other COPD’-associated deaths occurred among former workers and workers employed before the company implemented interventions reducing diacetyl exposure in 2001.

Conclusion

Workers at the microwave popcorn company experienced normal rates of all-cause mortality but higher rates of COPD-associated mortality, especially workers employed before the company reduced diacetyl exposure. The demonstrated excess in COPD-associated mortality suggests continued efforts to lower flavoring exposure are prudent.     

Alpha2-adrenergic regulation of NO production alters postoperative intestinal smooth muscle dysfunction in rodents

Alpha2-adrenergic receptor activation plays an important role in the development of postoperative ileus. Alpha2-adrenergic receptors also regulate nitric oxide (NO) production by the mononuclear phagocyte system. We have previously shown that intestinal manipulation leads to a significant increase in NO production by infiltrating monocytes within the intestinal muscularis. The purpose of this study was to investigate whether alpha2-adrenergic blockade with yohimbine would alter postsurgical intestinal smooth muscle dysfunction and NO production by infiltrating monocytes and macrophages within the intestinal muscularis. Rats underwent small bowel intestinal manipulation with or without yohimbine. In vivo gastrointestinal transit and in vitro jejunal circular muscle contractility was measured 24 h postoperatively. RT-PCR was used to detect inducible NO synthase (iNOS) expression. NO levels in tissue culture supernatants were measured. Immunohistochemistry was used to localize alpha2-adrenergic receptor expression in the intestinal muscularis. Yohimbine significantly decreased manipulation-induced delay in gastrointestinal transit and reversed the postoperative decrease in intestinal muscle contractility. Intestinal manipulation resulted in significant iNOS mRNA induction in the intestinal muscularis, which was markedly attenuated after yohimbine treatment. Yohimbine also significantly decreased the postoperative increase in NO released into intestinal muscularis tissue culture supernatant. Immunohistochemistry identified alpha2-adrenergic receptors on monocytes recruited postoperatively into the intestinal muscularis. This study demonstrates that alpha2-adrenergic receptor stimulation of the inflamed postoperative intestinal muscularis plays a significant role in aggravating postoperative ileus through an enhanced induction of iNOS mRNA and increased release of NO from manipulated intestinal muscularis.     

Individual variation in pup vocalizations and absence of behavioral signs of maternal vocal recognition in Weddell seals (Leptonychotes weddellii)

Individually stereotyped vocalizations often play an important role in relocation of offspring in gregarious breeders. In phocids, mothers often alternate between foraging at sea and attending their pup. Pup calls are individually distinctive in various phocid species. However, experimental evidence for maternal recognition is rare. In this study, we recorded Weddell seal (Leptonychotes weddellii) pup vocalizations at two whelping patches in Atka Bay, Antarctica, and explored individual vocal variation based on eight vocal parameters. Overall, 58% of calls were correctly classified according to individual. For males (n= 12) and females (n= 9), respectively, nine and seven individuals were correctly identified based on vocal parameters. To investigate whether mothers respond differently to calls of familiar vs. unfamiliar pups, we conducted playback experiments with 21 mothers. Maternal responses did not differ between playbacks of own, familiar, and unfamiliar pup calls. We suggest that Weddell seal pup calls may need to contain only a critical amount of individually distinct information because mothers and pups use a combination of sensory modalities for identification. However, it cannot be excluded that pup developmental factors and differing environmental factors between colonies affect pup acoustic behavior and the role of acoustic cues in the relocation process.     

Immunogenetic factors in beryllium sensitization and chronic beryllium disease

Exposure to beryllium in the workplace can cause beryllium sensitization and chronic beryllium disease. Sensitization to beryllium can be detected in the laboratory using the beryllium lymphocyte proliferation test. It was shown that anti-HLA antibodies could block the beryllium-specific response in the beryllium lymphocyte proliferation test, thereby implicating HLA genes in chronic beryllium disease. A supratypic genetic marker, HLA-DPB1*E69, was found to be strongly associated with immunologic sensitization to beryllium and chronic beryllium disease in beryllium workers. However, there are 40 HLA-DPB1 gene variants that have E69 but that also have other DNA sequence variations. The purpose of the study was to evaluate the evidence for potential differential susceptibility that may be associated with the physical characteristics of HLA protein molecules for which different HLA-DPB1*E69 variants code; that is, do some HLA-DPB1*E69 variants convey higher risk of beryllium sensitization and chronic beryllium disease than others. To do this, two approaches were pursued: first, detailed analysis of the findings from the published literature was performed, and second, computational chemistry was used to seek clues concerning the physical properties of the HLA protein molecules for which these alleles code. Among the 40 HLA-DPB1 gene variants that code for E69, molecular epidemiological studies have suggested a risk hierarchy, where some variants appear to convey low to moderate risk of chronic beryllium disease (e.g., HLA-DPB1*0201, approximately 3-fold increased risk), some convey an intermediate risk (e.g., HLA-DPB1*1901, approximately 5-fold) and others convey high risk (e.g., HLA-DPB1*1701, >10-fold). Molecular modeling has been used to further investigate a potential mechanistic basis for these observations. We found a strong correlation between the hierarchical order of risk of chronic beryllium disease associated with specific alleles and the predicted surface electrostatic potential and charge of the corresponding isotypes. Therefore, when alleles were grouped by the relative negative charge on the molecules for which they code, the data suggest that those alleles associated with the most negatively charged proteins carry the greatest risk of beryllium sensitization and disease.     

Morphology of the prothoracic discs and associated sensilla of Acanthocnemus nigricans (Coleoptera, Acanthocnemidae)

The Australian ‘little ash beetle’ Acanthocnemus nigricans (Coleoptera, Cleroidea, Acanthocnemidae) is attracted by forest fires. A. nigricans has one pair of unique prothoracic sensory organs and it has been speculated that these organs may play a role in fire detection. Each organ consists of a cuticular disc, which is fixed over an air-filled cavity. On the outer surface of the disc, about 90 tiny cuticular sensilla are situated. The poreless outer peg of a sensillum is 3–5 μm long and is surrounded by a cuticular wall. One ciliary sensory cell innervates the peg. As a special feature, the outer dendritic segment is very short already terminating below the cuticle. A massive electron-dense cylindrical rod, which most probably represents the hypertrophied dendritic sheath, extends through the cuticular canal connecting the tip of the outer dendritic segment to the peg. The dendritic inner segment and the soma are fused indistinguishably. Thin, leaflike extensions of glial cells deeply extend into that conjoint and considerably enlarged compartment which also contains large numbers of mitochondria. In summary, the sensilla of the sensory disc of A. nigricans represent a new type of insect sensillum of hitherto unknown function. The possible role of the prothoracic sensory organ in fire detection is discussed.     

Photochemistry of dyes and fluorochromes used in biology and medicine: some physicochemical background and current applications

An overview of the basic principles of photochemistry is presented to facilitate discussion of fluorescence, quenching and quantum yields. These topics in turn provide the foundation for an account of fluorescence spectroscopy and its application to microscopy. A brief overview of light microscopy and the application of fluorescence microscopy is given. The influences of molecular features, such as aromatic character and substitution patterns, on color and fluorescence are described. The concept of color fading is considered with particular reference to its effect on microscopic preparations. A survey of representative fluorescent probes is provided, and their sensitivity, application, and limitations are described. The phototoxicity of fluorescent molecules is discussed using biomembranes and DNA as examples of targets of toxicity. Photodynamic therapy, a relatively new clinical application of phototoxicity, is described. Both anticancer and antimicrobial applications are noted, and an assessment is given of the current ideas on the ideal physicochemical properties of the sensitizing agents for such applications.