World Trade Center exposure, post-traumatic stress disorder, and subjective cognitive concerns in a cohort of rescue/recovery workers.

OBJECTIVE: To determine whether World Trade Center (WTC)-exposure intensity and post-traumatic stress disorder (PTSD) are associated with subjective cognitive change in rescue/recovery workers. METHOD: The population included 7875 rescue/recovery workers who completed a subjective cognition measure, the Cognitive Function Instrument (CFI), between 3/1/2018 and 2/28/2019 during routine monitoring, indicating whether they had experienced cognitive and functional difficulties in the past year. Higher scores indicated greater self-perceived cognitive change. Probable PTSD, depression, and alcohol abuse were evaluated by validated mental health screeners. Logistic regression assessed the associations of WTC exposure and current PTSD with top-quartile (≥2) CFI score, and of early post-9/11 PTSD with top-quartile CFI in a subpopulation (N = 6440). Models included demographics, smoking, depression, and alcohol abuse as covariates. RESULTS: Mean age at CFI completion was 56.7 ± 7.7 (range: 36-81). Participants with high-intensity WTC exposure had an increased likelihood of top-quartile CFI score (odds ratio[OR] vs. low exposure: 1.32, 95%CI: 1.07-1.64), controlling for covariates. Current and early PTSD were both associated with top-quartile CFI (OR: 3.25, 95%CI: 2.53-4.19 and OR: 1.56, 95%CI: 1.26-1.93) respectively. CONCLUSIONS: High-intensity WTC exposure was associated with self-reported cognitive change 17 years later in rescue/recovery workers, as was PTSD. Highly WTC-exposed subgroups may benefit from additional cognitive evaluation and monitoring of cognition over time.

Effect of smoking cessation on cough reflex sensitivity.

Recent studies have shown that cigarette smokers have diminished cough reflex sensitivity compared with nonsmokers. The current authors proposed a mechanism of chronic cigarette smoke-induced desensitisation of airway cough receptors. To investigate this hypothesis, cough sensitivity to inhaled capsaicin (C5) in chronic smokers was measured both while they were actively smoking and 2, 6, 12 and 24 weeks after smoking cessation. In total, 29 subjects underwent baseline capsaicin challenge while smoking and 2 weeks after smoking cessation. Mean+/-sem log C5 fell from 1.86+/-0.12 to 1.60+/-0.12, demonstrating significant enhancement of cough reflex sensitivity. Of the total, 20, 18 and 14 subjects successfully abstained from smoking for 6, 12 and 24 weeks, respectively. Mean log C5 values after 12 and 24 weeks of smoking cessation were significantly diminished from baseline. In a control group of smokers, mean log C5 did not decrease from baseline after 6, 12 and 24 weeks. Overall, the log C5 profile of the smoking cessation group showed a clear, linearly decreasing trend over time compared with the control group. Even after many years of smoking, cough sensitivity is enhanced as early as 2 weeks after smoking cessation. Given the importance of an intact cough reflex, these changes may provide clinical benefit.

An evaluation of Open Airways for Schools: using college students as instructors.

Fifteen years ago, Open Airways for Schools (OAS) was found to be an effective asthma education program for elementary school children when taught by professionals. To determine whether OAS is effective when taught by college students and whether it could withstand potential cohort effects, 54 inner-city fourth and fifth graders were taught OAS. Paired t-tests revealed that OAS improved asthma knowledge, self-efficacy, self-management skills, social support, and perception of well-being (p < 0.05). Fifteen years later OAS continues to improve children's self-management skills. Facilitators with little prior experience who received brief training in asthma knowledge and group leadership skills can effectively teach OAS.

A P450 BM-3 mutant hydroxylates alkanes, cycloalkanes, arenes and heteroarenes.

P450 monooxygenases from microorganisms, similar to those of eukaryotic mitochondria, display a rather narrow substrate specificity. For native P450 BM-3, no other substrates than fatty acids or an indolyl-fatty acid derivative have been reported (Li, Q.S., Schwaneberg, U., Fischer, P., Schmid, R.D., 2000. Directed evolution of the fatty-acid hydroxylase P450BM-3 into an indole-hydroxylating catalyst. Chem. Eur. J. 6 (9), 1531-1536). Engineering the substrate specificity of Bacillus megaterium cytochrome P-450 BM3: hydroxylation of alkyl trimethylammonium compounds. Biochem. J. 327, 537-544). We thus were quite surprised to observe, in the course of our investigations on the rational evolution of this enzyme towards mutants, capable of hydroxylating shorter-chain fatty acids, that a triple mutant P450 BM-3 (Phe87Val, Leu188-Gln, Ala74Gly, BM-3 mutant) could efficiently hydroxylate indole, leading to the formation of indigo and indirubin (Li, Q.S., Schwaneberg, U., Fischer, P., Schmid, R.D., 2000. Directed evolution of the fatty-acid hydroxylase P450BM-3 into an indole-hydroxylating catalyst. Chem. Eur. J. 6 (9), 1531-1536). Indole is not oxidized by the wild-type enzyme; it lacks the carboxylate group by which the proper fatty acid substrates are supposed to be bound at the active site of the native enzyme, via hydrogen bonds to the charged amino acid residues Arg47 and Tyr51. Our attempts to predict the putative binding mode of indole to P450 BM-3 or the triple mutant by molecular dynamics simulations did not provide any useful clue. Encouraged by the unexpected activity of the triple mutant towards indole, we investigated in a preliminary, but systematic manner several alkanes, alicyclic, aromatic, and heterocyclic compounds, all of which are unaffected by the native enzyme, for their potential as substrates. We here report that this triple mutant indeed is capable to hydroxylate a respectable range of other substrates, all of which bear little or no resemblance to the fatty acid substrates of the native enzyme.

P450 in biotechnology: zinc driven omega-hydroxylation of p-nitrophenoxydodecanoic acid using P450 BM-3 F87A as a catalyst.

Cytochrome P450 enzymes require the delivery of two electrons to the heme protein for their enzymatic function. NADPH or NADH are usually used as reduction equivalents. In the absence of a substrate, NADPH may inactivate P450 enzymes. Furthermore, it is expensive, making it unsuitable for the preparative synthesis of fine chemicals. Approaches for replacing NADPH with an electrochemically generated reduction by using platinum-electrodes and different mediators are known. In the present study, NADPH was substituted by the mediator cobalt(III)sepulchrate and zinc dust that serves as an electron source. The mutated fatty acid hydroxylase P450 BM-3 F87A from Bacillus megaterium was chosen as a catalyst, since it shows a three-fold higher sensitivity and a nearly five-fold higher activity for p-nitrophenoxydodecanoic acid (12-pNCA) than the wild-type enzyme. The formation of p-nitrophenolate can easily be monitored using a photometer at 410 nm. The turnover rate of the zinc/cobalt(III)sepulchrate system reaches 20% of the NADPH activity. Compared to the electrochemical approaches the activity is at least 77% higher (turnover 125 eq min-1). The presented alternative cofactor system can be used instead of NADPH or expensive electrochemical devices (platinum electrodes) for fine chemical synthesis.

Visual and Infrared Assessment of Root Colonization of Apple Trees by Phymatotrichopsis omnivora.

Root systems of 5-year-old, trellised apple trees (Malus domestica Borkh) on cv. M.7a root-stocks were assessed for the presence of fungal strands of Phymatotrichopsis omnivora (Duggar) Hennebert in two orchards in central Texas. Fungal advance within each orchard was assessed in five directions. Pathogen growth (P < 0.01) occurred beyond symptomatic trees along and perpendicularly across rows. In one orchard, 80% of the first asymptomatic trees were infected along rows, followed by 60% infection perpendicularly across rows. In the other orchard, there was 100% infection of the first asymptomatic trees along rows and 60% infection perpendicularly across rows. No growth was observed diagonally across rows in either orchard. Infrared readings of canopy temperature and differences between canopy temperature and air temperature were significant (P < 0.01) for predicting infection of asymptomatic, infected trees in one orchard. Trees were shown to have extensive taproot decay and infection of lateral roots before canopy symptoms began to develop. Root diameter appeared to have no effect on the growth of the fungus.

A high-density integrated genetic linkage and radiation hybrid map of the laboratory rat.

The laboratory rat (Rattus norvegicus) is a key animal model for biomedical research. However, the genetic infrastructure required for connecting phenotype and genotype in the rat is currently incomplete. Here, we report the construction and integration of two genomic maps: a dense genetic linkage map of the rat and the first radiation hybrid (RH) map of the rat. The genetic map was constructed in two F2 intercrosses (SHRSP x BN and FHH x ACI), containing a total of 4736 simple sequence length polymorphism (SSLP) markers. Allele sizes for 4328 of the genetic markers were characterized in 48 of the most commonly used inbred strains. The RH map is a lod >/= 3 framework map, including 983 SSLPs, thereby allowing integration with markers on various genetic maps and with markers mapped on the RH panel. Together, the maps provide an integrated reference to >3000 genes and ESTs and >8500 genetic markers (5211 of our SSLPs and >3500 SSLPs developed by other groups). [Bihoreau et al. (1997); James and Tanigami, RHdb (http:www.ebi.ac.uk/RHdb/index.html); Wilder (http://www.nih.gov/niams/scientific/ratgbase); Serikawa et al. (1992); RATMAP server (http://ratmap.gen.gu.se)] RH maps (v. 2.0) have been posted on our web sites at http://goliath.ifrc.mcw.edu/LGR/index.html or http://curatools.curagen.com/ratmap. Both web sites provide an RH mapping server where investigators can localize their own RH vectors relative to this map. The raw data have been deposited in the RHdb database. Taken together, these maps provide the basic tools for rat genomics. The RH map provides the means to rapidly localize genetic markers, genes, and ESTs within the rat genome. These maps provide the basic tools for rat genomics. They will facilitate studies of multifactorial disease and functional genomics, allow construction of physical maps, and provide a scaffold for both directed and large-scale sequencing efforts and comparative genomics in this important experimental organism.

Using Airborne Digital Imagery for Detecting Oak Wilt Disease.

Color-infrared (CIR) digital imagery was evaluated as a remote sensing tool for detecting oak wilt disease in live oak (Quercus fusiformis). Aerial CIR digital imagery and CIR photography were obtained concurrently of a live oak forested area in south-central Texas affected by oak wilt. Dead, diseased, and healthy live oak trees could generally be delineated as well in the digital imagery as in the CIR photography. Light reflectance measurements obtained in the field showed that dead, diseased, and healthy trees had different visible and near-infrared reflectance values.

Glycolipid-independent sorting of a secretory glycoprotein to the apical surface of polarized epithelial cells.

Proteins attached to the membrane by a glycosylphosphatidylinositol (GPI)-anchor cluster together with glycolipids in detergent-insoluble complexes at the site of sorting in the trans-Golgi network. This process has been shown to be critical for the targeting of these proteins to the apical cell surface in polarized epithelial cells. We show in this study that gp80 (clusterin), an apically secreted glycoprotein, is not included in detergent-insoluble complexes in Madin-Darby canine kidney cells. Furthermore in Fisher rat thyroid cells, which target GPI-anchored proteins preferentially to the basolateral cell surface, gp80 is secreted apically. Together these results suggest that this secretory glycoprotein and GPI-linked proteins use different mechanisms to reach the apical membrane.

Sorting of gp80 (GPIII, clusterin), a marker protein for constitutive apical secretion in Madin-Darby canine kidney (MDCK) cells, into the regulated pathway in the pheochromocytoma cell line PC12.

We have studied the biogenesis and transport of a marker protein for constitutive apical secretion in Madin-Darby canine kidney (MDCK) cells, the gp80 glycoprotein complex (clusterin, apolipoprotein J, complement lysis inhibitor), in the rat pheochromocytoma cell line, PC12, by pulse-chase analysis of the endogeneously expressed complex. We demonstrate that in PC12 cells, gp80 is secreted via the regulated pathway, although sorting into that pathway is inefficient. Of the newly synthesized complex, 50% is released constitutively during a 2.5 h chase, 15% is released by depolarization-induced secretion, 35% stay associated with the cells. In contrast to their pivotal role in the constitutive apical exocytosis of the gp80 complex in MDCK cells, the N-linked carbohydrate moieties are dispensible for the sorting of this protein into the regulated pathway in PC12 cells, suggesting that distinct signals and sorting mechanisms are involved in these pathways.

Relationships among pathogenic and nonpathogenic isolates of Fusarium oxysporum based on the partial sequence of the intergenic spacer region of the ribosomal DNA.

Using PCR, we amplified and sequenced approximately 1,000 bp of the 5' end of the intergenic spacer (IGS) of the rDNA in 15 isolates of Fusarium oxysporum and one isolate of F. subglutinans. Isolates were selected to represent diversity in our collection based on differences in pathogenic race, vegetative compatibility group (VCG), mitochondrial DNA (mtDNA) haplotype, IGS haplotype, and DNA fingerprint. The objective of this research was to clarify the origin of virulence within F. oxysporum, the relationship between pathogenic and nonpathogenic strains, and the evolution of the different races of F. oxysporum f. sp. melonis. Bootstrapped parsimony analysis of the partial IGS sequence data identified a phylogenetic tree with highly significant branches. The two F. oxysporum f. sp. melonis VCGs, 0131 and 0134, were separated into distinct lineages. Race was not distinguished by significant IGS sequence differences within the pathogen VCGs. One exception was a race 1 isolate which was associated with VCG 0131 but, based on both mtDNA and IGS haplotype, had greater affinity with VCG 0134. Two IGS sequence types were found in this race 1 isolate, one suggesting an affiliation with VCG 0131 and the other similar to isolates in VCG 0134. This may have resulted from past somatic or sexual interactions between F. oxysporum f. sp. melonis, VCGs 0131 and 0134. Nonpathogens that were vegetatively compatible with the pathogen were not closely related to the pathogen based on IGS sequence data. Thus, nonpathogens and pathogens may share common alleles at vegetative compatibility loci by coincidence rather than because of recent clonal derivation from a common ancestor.

Expression of mRNA for vasoactive intestinal peptide in rat small intestine.

Transplantation of small intestine is a neural model that permits studies of expression of the neuropeptide, vasoactive intestinal peptide, following extrinsic denervation, transection of intrinsic neural pathways, and an ischemic interval. Tissue levels of vasoactive intestinal peptide were examined at 3 months in ileum from a sham operation, in ileum after resection of proximal small intestine, in ileum after resection of proximal small intestine and extrinsic denervation, in ileum after resection of proximal small intestine and 30 min of ischemia, and in ileum obtained 3 months after ileal isografting in Lewis-to-Lewis combinations. Vasoactive intestinal peptide levels were increased in transplanted rat ileum, resection controls, denervation controls, and ischemic controls compared to sham-operated ileum (pANOVA < 0.01). The increased levels of this peptide were highest in denervation controls and lowest in ischemic controls. Northern blot analysis using rat vasoactive intestinal peptide cDNA identified a single 1.7-kb transcript in normal and transplanted rat ileum. The density of vasoactive intestinal peptide transcripts was increased in transplanted ileum (8450 +/- 540) compared to normal ileum (5790 +/- 620) (P < 0.01), and the ratio of this transcript to glyceraldehyde-3-phosphate dehydrogenase density units was also increased in transplanted ileum (0.81 +/- 0.08) compared to normal ileum (0.40 +/- 0.07; P < 0.01). Enhanced transcriptional regulation was the likely mechanism for increased tissue vasoactive intestinal peptide. The increased tissue levels appeared to be a response to extrinsic denervation and transection of intrinsic neural pathways, while an ischemic interval appeared to decrease tissue levels of the peptide.

Intraspecific variation within populations of Fusarium oxysporum based on RFLP analysis of the intergenic spacer region of the rDNA.

Fifty-six isolates of Fusarium oxysporum, including F. oxysporum f. sp. melonis and nonpathogenic strains, were chosen from a larger collection to represent diversity in vegetative compatibility groups (VCGs), mitochondrial DNA (mtDNA) haplotype, geographic distribution, and virulence. Using PCR, a 2.6-kb fragment including the intergenic spacer (IGS) region of the ribosomal DNA was amplified from each isolate. The enzymes EcoRI, Sau3A, CfoI, and AvaII, cut this fragment differentially, revealing 5, 6, 6, and 7 patterns, respectively. Among the 56 isolates, a total of 13 unique IGS haplotypes was identified. Among most F. o. melonis isolates. IGS haplotype correlated with VCG and mtDNA haplotype, but did not differentiate among races. However, a race 1 isolate found in VCG 0131 shared virulence, mtDNA, and IGS haplotypes characteristic of VCG 0134; this isolate may represent a conversion in VCG from 0134 to 0131. Four nonpathogens shared the pathogen vegetative compatibility phenotypes. One race 1, 2 isolate associated with VCG 0134 shared both IGS haplotype and VCG with a nonpathogen, but these isolates did not share the same mtDNA haplotype. Another nonpathogenic isolate shared mtDNA and IGS haplotypes with pathogen group 0131 and may simply be an avirulent mutant of a pathogenic strain. For the other two nonpathogenic isolates, vegetative compatibility indicated a close relationship to the pathogen, but differences in both mtDNA and IGS haplotype suggest otherwise. Overall, the IGS haplotype was more variable among the nonpathogenic F. oxysporum VCGs among which 12 of the 13 IGS haplotypes were found. Nonpathogenic isolates that shared a common mtDNA haplotype, but were associated with different VCGs, often had different IGS haplotypes.

RFLPs in mitochondrial and nuclear DNA indicate low levels of genetic diversity in the oak wilt pathogen Ceratocystis fagacearum.

Genetic diversity in the oak wilt pathogen Ceratocystis fagacearum was assessed using restriction fragment length polymorphisms (RFLPs) of the mitochondrial DNA (mtDNA) and anonymous RFLP loci in the nuclear DNA (nuDNA). No genetic variation was detected in the mtDNA among 27 isolates sampled from a broad geographical area. Southern hybridization to 100 anonymous, random, nuDNA probes detected a low level of variation among nine of the isolates. Only 35 out of 437 probe-enzyme combinations detected RFLPs. Most of the RFLPs appeared to result from insertions and deletions of less than 200 bp. A composite multilocus haplotype based on hybridization to six anonymous probes could differentiate each of the nine isolates tested, suggesting that these probes may be useful for further studies of the population biology and epidemiology of this pathogen. Hypotheses are presented to account for the low level of genetic variation.

The oak wilt enigma: perspectives from the Texas epidemic.

Ceratocystis fagacearum (Bretz) Hunt, the oak wilt pathogen, is currently causing massivel osses of semievergreen live oaks (Quercus fusiformis Small and Q. virginiana Mill.) in central Texas. Given the relatively limited oak mortality caused by C. fagacearum in the deciduous forests of the North Central, Midwestern, and Mid-Atlantic United States, this Texas epidemic was not anticipated. The intensity of oak wilt in Texas is attributed to a number of factors related to host characteristics and the ability of the pathogen to adapt to limiting environmental conditions. Oak wilt management in semievergreen oaks requires considerable revision of the control techniques previously designed for deciduous oaks. The Texas oak wilt epidemic provides a new perspective from which to evaluate questions concerning oak wilt, including the origins of the pathogen as well as the potential for future losses in unaffected oak forests.

Sorting of a secretory protein (gp80) to the apical surface of Caco-2 cells.

We have investigated the synthesis and polarized secretion of the exogenous gp80 glycoprotein complex in the human epithelial adenocarcinoma cell line, Caco-2. gp80 is secreted at the apical surface of Madin-Darby canine kidney (MDCK) cells and should, therefore, display the signal(s) required for sorting into the apical exocytic pathway. In Caco-2 cells, no bona fide secretory protein released preferentially at the apical surface has been described so far. To address the question of whether Caco-2 cells possess a machinery capable of delivery of secretory proteins at the apical surface, we stably transfected the cells with a recombinant gene coding for the gp80 glycoprotein complex. Pulse-chase analysis showed that stably transfected Caco-2 cells secrete gp80 quantitatively into the medium. In polarized layers of filter-grown Caco-2 cells, the protein was secreted predominantly at the apical surface, demonstrating the ability of the cells to efficiently sort secretory proteins directly into the apical exocytic pathway. Our results further demonstrate that the apical targeting information of gp80 recognized by MDCK cells is also recognized by Caco-2 cells.

Health care costs in workers' compensation.

Although the costs of both the medical and indemnity components of workers' compensation have increased substantially in the past several years, the costs of medical benefits grew far more dramatically.

Molecular cloning of gp 80, a glycoprotein complex secreted by kidney cells in vitro and in vivo. A link to the reproductive system and to the complement cascade.

cDNA clones coding for the gp 80 heterodimeric glycoprotein complex secreted constitutively at the apical surface of Madin-Darby canine kidney (MDCK) cells have been isolated from MDCK cDNA libraries in lambda gt11 and lambda gt10. The cloned sequences encode a polypeptide chain of 445 amino acids. The deduced amino acid sequence of the gp 80 protein reveals 80% homology to rat SGP-2, a major secretory protein of the testes epithelium and 83% homology to SP-40,40, a human complement-associated protein. SGP-2 and SP-40,40 have been proposed to be serum and seminal forms of the same protein. The sequence homology as well as the results of Southern and Northern blot analyses and immunological studies suggest that gp 80 is the canine homolog of the rat SGP-2 and the human SP-40,40. The protein is expressed in the embryonic kidney already early during organogenesis. In the adult kidney the protein has been localized along the luminal surfaces of the proximal and distal tubule and the collecting duct cells.