We have compared the use of five nonvaccine antigens to the use of conventional vaccine antigens, pertussis toxin (PT), and filamentous hemagglutinin (FHA) for the serological diagnosis of pertussis by enzyme-linked immunosorbent assay (ELISA). The nonvaccine antigens included the catalytic region of adenylate cyclase toxin (CatACT), the C-terminal region of FHA (C-FHA), lipooligosaccharide (LOS), the peptidoglycan-associated lipoprotein (PAL), and the BrkA protein. The serological responses of individuals with culture-confirmed pertussis were compared to those of adults with no recent history of a coughing disease. An immunoglobulin G (IgG) ELISA for PT was the most sensitive (92.2%) test for the serodiagnosis of pertussis. Of the nonvaccine antigens, ELISA for IgG responses to CatACT (sensitivity, 62.8%), C-FHA (sensitivity, 39.2%), and LOS IgA (sensitivity, 29.4%) were less sensitive but could also distinguish culture-positive individuals from control individuals. The use of a combination of multiple ELISA targets improved the sensitivity of the assay for serological diagnosis. Elevated IgG and IgA antibody titers persisted for more than a year in the individuals with culture-confirmed pertussis.

Biosynthesis of caffeoylquinic acids occurs via the phenylpropanoid pathway in which the phenylalanine ammonia-lyase (<em>PAL</em>) acts as a key-control enzyme. A full-length cDNA (pF6), corresponding to a <em>PAL</em> gene (Cc<em>PAL</em>1), was isolated by screening a Coffea canephora fruit cDNA library and its corresponding genomic sequence was characterized. Amplification of total DNA from seven Coffea species revealed differences in intronic length. This interspecific polymorphism was used to locate the gene on a genetic map established for a backcross progeny between Coffea pseudozanguebariae and C. dewevrei. The Cc<em>PAL</em>1 gene was found on the same linkage group, but genetically independent, as a caffeoyl-coenzyme A-O-methyltransferase gene, another gene intervening in the phenylpropanoid pathway. In the same backcross, a lower caffeoylquinic acid content was observed in seeds harvested from plants harbouring the C. pseudozanguebariae Cc<em>PAL</em>1 allele. Involvement of the Cc<em>PAL</em>1 allelic form in the differential accumulation of caffeoylquinic acids in coffee green beans is then discussed.

A novel peptidoglycan-associated lipoprotein (PAL) was found in the cell envelope of Proteus mirabilis. This protein showed the following properties: (1) The apparent molecular weight in sodium dodecyl sulfate (SDS) polyacrylamide gel was 18,000. (2) The protein was present in the cell envelope in a form very closely, but not covalently, associated with the peptidoglycan layer. (3) The protein was recovered predominantly from the outer membrane fraction after separation of the cell envelopes. (4) [1-14C]Palmitic acid and [2-3H]glycerol were incorporated into the protein. (5) The protein contained covalently linked fatty acids (about 3 mol of fatty acid per mol of protein). (6) An unidentified compound was present in the hydrolysate of the protein. These properties, except for molecular weight and non-covalent association with the peptidoglycan, showed resemblance to those of Braun's lipoprotein. However the protein was distinct from Braun's lipoprotein in regard to amino acid composition. A similar peptidoglycan-associated lipoprotein (PAL) was present widely in the cell envelopes of various Gram-negative bacteria. P. mirabilis contains about twelve times as much PAL as Escherichia coli. Antiserum against PAL of P. mirabilis was cross-reactive against PAL of E. coli, but not against Braun's lipoprotein of E. coli.

Light and fungal elicitor induce mRNA encoding 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase in suspension cultured cells of parsley (Petroselinum crispum L.). The kinetics and dose response of mRNA accumulation were similar for DAHP synthase and phenylalanine ammonia-lyase (PAL). Six micrograms of elicitor from Phytophthora megasperma f. glycinia gave a detectable induction within 1 hour. Induction of DAHP synthase and PAL mRNAs by light was transient, reaching maximal levels at 4 hours and returning to pretreatment levels after 24 hours. Our data suggest that either light or fungal elicitor transcriptionally activate DAHP synthase. A coordinate regulation for key enzymes in the synthesis of primary and secondary metabolites is indicated.

A study of the expression of a bean phenylalanine ammonia-lyase (<em>PAL</em>) promoter/beta-glucuronidase gene fusion in transgenic tobacco has shown that the <em>PAL</em>2 promoter has a modular organization. Expression of the <em>PAL</em>2 promoter in the vascular system involves positive and negative regulatory cis elements. Among these elements is an AC-rich motif implicated in xylem expression and a suppressing cis element for phloem expression. Using radiolabelled complementary oligonucleotides bearing the AC-rich motif, a cDNA clone encoding a DNA-binding protein has been isolated from a tobacco lambda gt11 expression library. This factor, named AC-rich binding factor (ACBF), showed binding specificity to the AC-rich region. The specificity of ACBF for the AC-rich region was also shown using a gel retardation assay with an ACBF recombinant protein extract. The deduced amino acid sequence from ACBF contains a long repeat of glutamine residues as found in well characterized transcription factors. Interestingly, ACBF shared sequence similarity to conserved amino acid motifs found in RNA-binding proteins. Genomic gel blot analysis indicated the presence of a small gene family of sequences related to ACBF within the tobacco nuclear genome. Analysis of tobacco mRNA using the ACBF cDNA as probe showed that while ACBF mRNA was present in all tissues examined, the highest transcript accumulation occurred in stem tissues. The functional characteristics of the AC-rich sequence were examined in transgenic tobacco. A heptamer of the AC-rich sequence, in front of a minimal 35S promoter from cauliflower mosaic virus (-46 to +4), conferred specific expression in xylem.

A survey was conducted on the present behavioral characteristics of 187 cases of adult autism in patients over 18 years of age employing Achenbach's Child Behavior Checklist (CBCL). When their behavioral characteristics were evaluated in relation to Present Language Developmental Level (PLDL) and Present Adaptive Level (PAL), it was seen that greater variation in behavior characteristics was seen among those exhibiting increasingly lower PLDL and PAL scores. Behavior characteristics reminiscent of depression were noted even among those exhibiting high PLDL. Behavior pointing to obsession was found in common among almost all cases of autism irrespective of PLDL or PAL. Psychotic symptoms such as hallucinations and delusions were absent in most cases. The results of the present study were indicative not only of the significance of obsessive behavior in autism, but also its significance in terms of delving further into the psychopathology of the disorder.

BACKGROUND

Periodontitis is a chronic infective disease of the gums caused by bacteria present in dental plaque. This condition induces the breakdown of the tooth supporting apparatus until teeth are lost. Surgery may be indicated to arrest disease progression and regenerate lost tissues. Several surgical techniques have been developed to regenerate periodontal tissues including guided tissue regeneration (GTR), bone grafting (BG) and the use of enamel matrix derivative (EMD). EMD is an extract of enamel matrix and contains amelogenins of various molecular weights. Amelogenins are involved in the formation of enamel and periodontal attachment formation during tooth development.

OBJECTIVE

To test whether EMD is effective, and to compare EMD versus GTR, and various BG procedures for the treatment of intrabony defects.

METHODS

We searched the Cochrane OHG Trials Register, Cochrane Central Register of Controlled Trials, MEDLINE and EMBASE. Several journals were handsearched. No language restrictions were applied. Authors of RCTs identified, personal contacts and the manufacturer were contacted to identify unpublished trials. Most recent search: May 2005.

METHODS

RCTs on patients affected by periodontitis having intrabony defects of at least 3 mm treated with EMD compared with open flap debridement, GTR and various BG procedures with at least 1 year follow up. The outcome measures considered were: tooth loss, changes in probing attachment levels (PAL), pocket depths (PPD), gingival recessions (REC), bone levels from the bottom of the defects on intraoral radiographs, aesthetics and adverse events. The following time-points were to be evaluated: 1, 5 and 10 years.

METHODS

Screening of eligible studies, assessment of the methodological quality of the trials and data extraction were conducted in duplicate and independently by two authors. Results were expressed as random-effects models using mean differences for continuous outcomes and risk ratios (RR) for dichotomous outcomes with 95% confidence intervals (CI). It was decided not to investigate heterogeneity, but a sensitivity analysis for the risk of bias of the trials was performed.

RESULTS

Ten trials were included out of 29 potentially eligible trials. No included trial presented data after 5 years of follow up, therefore all data refer to the 1-year time point. A meta-analysis including eight trials showed that EMD treated sites displayed statistically significant PAL improvements (mean difference 1.2 mm, 95% CI 0.7 to 1.7) and PPD reduction (0.8 mm, 95% CI 0.5 to 1.0) when compared to placebo or control treated sites, though a high degree of heterogeneity was found. Significantly more sites had < 2 mm PAL gain in the control group, with RR 0.48 (95% CI 0.29 to 0.80). Approximately six patients needed to be treated (NNT) to have one patient gaining 2 mm or more PAL over the control group, based on a prevalence in the control group of 35%. No differences in tooth loss or aesthetic appearance as judged by the patients were observed. When evaluating the only two trials at a low risk of bias in a sensitivity analysis, the effect size for PAL was 0.6 mm, which was less than 1.2 mm for the overall result. Comparing EMD with GTR (five trials), GTR showed a statistically significant increase of REC (0.4 mm) and significantly more postoperative complications. No trials were found comparing EMD with BG.

CONCLUSIONS

One year after its application, EMD significantly improved PAL levels (1.2 mm) and PPD reduction (0.8 mm) when compared to a placebo or control, however, the high degree of heterogeneity observed among trials suggests that results have to be interpreted with great caution. In addition a sensitivity analyses indicated that the overall treatment effect might be overestimated. The actual clinical advantages of using EMD are unknown. With the exception of significantly more postoperative complications in the GTR group, there was no evidence of clinically important differences between GTR and EMD.

Mediator complexes are large multiprotein assemblies that function in the regulation of eukaryotic gene transcription. In yeast, certain mediator subunits appear to comprise a subcomplex that acts in the regulation of a specific subset of genes. We investigated in a metazoan, Caenorhabditis elegans, the roles and interactions of two of those subunits, CeTRAP240/let-19 and CeTRAP230/dpy-22. We found that CeTRAP240/let-19 contains four domains that are conserved in the human TRAP240 protein and that one of those domains displays intrinsic transcriptional repression activity. Using RNA interference, we found that reduced expression of CeTRAP240/let-19 displayed a high penetrance of embryonic lethality in F1 progeny; animals that escaped embryonic arrest showed mutant phenotypes such as burst vulva and molting defects. CeTRAP240/let-19 appeared to affect specific genes, as CeTRAP240/let-19(RNAi) led to selectively reduced expression of a subset of reporter genes examined. Genetic experiments supported the view that CeTRAP240/let-19 and CeTRAP230/dpy-22, like their Drosophila and yeast counterparts, can operate on common pathways. Thus, a male tail phenotype caused by the pal-1(e2091) mutation was suppressed not only by CeTRAP230/dpy-22 mutants, as reported previously, but also by reduced expression of CeTRAP240/let-19. Additionally, CeTRAP240/let-19(RNAi) in a CeTRAP230/dpy-22 mutant background produced a strong synthetic lethal phenotype. Overall, our results establish specific roles of CeTRAP240/let-19 in C. elegans embryonic development and a functional interaction between CeTRAP240/let-19 and CeTRAP230/dpy-22. Interestingly, whereas this interaction has been conserved from yeast to mammals, the subcomplex modulates metazoan-specific genetic pathways, likely in addition to those also controlled in yeast.

The surprisingly high catalytic activity and selectivity of enzymes stem from their ability to both accelerate the target reaction and suppress competitive reaction pathways that may even be dominant in the absence of enzymes. For example, histidine and phenylalanine ammonia-lyases (HAL and PAL) trigger the abstraction of the nonacidic beta protons of these amino acids while leaving the much more acidic ammonium hydrogen atoms untouched. Both ammonia-lyases have a catalytically important electrophilic group, which was believed to be dehydroalanine for 30 years but has now been revealed by X-ray crystallography and UV spectroscopy to be a highly electrophilic 5-methylene-3,5-dihydroimidazol-4-one (MIO) group. Experiments suggest that the reaction is initiated by the electrophilic attack of MIO on the aromatic ring of the substrate. This incomplete Friedel-Crafts-type reaction leads to the activation of a beta proton and its stereospecific abstraction, followed by the elimination of ammonia and regeneration of the MIO group. The plausibility of such a mechanism is supported by a synthetic model. The application of the PAL reaction in the biocatalytic synthesis of enantiomerically pure alpha-amino beta-aryl propionates from aryl acrylates is also discussed.

Secreted peptides, produced by enzymatic processing of larger precursor molecules, are found throughout the animal kingdom and play important regulatory roles as neurotransmitters and hormones. Many require a carboxy-terminal modification, involving the conversion of a glycine residue into an α-amide, for their biological activity. Two sequential enzymatic activities catalyze this conversion: a monooxygenase (peptidylglycine α-hydroxylating monooxygenase or PHM) and an amidating lyase (peptidyl-α-hydroxyglycine α-amidating lyase or PAL). In vertebrates, these activities reside in a single polypeptide known as peptidylglycine α-amidating monooxygenase (PAM), which has been extensively studied in the context of neuropeptide modification. Bifunctional PAMs have been reported from some invertebrates, but the phylogenetic distribution of PAMs and their evolutionary relationship to PALs and PHMs is unclear. Here, we report sequence and expression data for two PAMs from the coral Acropora millepora (Anthozoa, Cnidaria), as well as providing a comprehensive survey of the available sequence data from other organisms. These analyses indicate that bifunctional PAMs predate the origins of the nervous and endocrine systems, consistent with the idea that within the Metazoa their ancestral function may have been to amidate epitheliopeptides. More surprisingly, the phylogenomic survey also revealed the presence of PAMs in green algae (but not in higher plants or fungi), implying that the bifunctional enzyme either predates the plant/animal divergence and has subsequently been lost in a number of lineages or perhaps that convergent evolution or lateral gene transfer has occurred. This finding is consistent with recent discoveries that other molecules once thought of as "neural" predate nervous systems.

An amine-catalyzed reaction has been discovered that couples alpha-allenic esters with N-acyl imines in good to excellent yields (up to 88%). Extension of this methodology from the study of achiral pyridine-based catalysis to chiral peptide-based scaffolds is presented. The approach culminated in the identification of a tetrameric peptide sequence containing an embedded pyridylalanine (Pal) residue as an efficient asymmetric catalyst for enantioselective coupling reactions. The unique allenic products are obtained with enantiomer ratios of up to approximately 95:5 (up to >98:2 following recrystallization).

We compared the self-reported frequency of recreational exercise and corresponding metabolic equivalent (MET)-minutes with physical activity measured with a position and motion sensor in pregnant women. One hundred and twelve women in the Norwegian Mother and Child Cohort Study (MoBa) completed questions about weekly participation in recreational exercise by week 17 of pregnancy and participated in the validation study around week 20. Data from a validated motion sensor (ActiReg) that measures physical activity and total energy expenditure (TEE) served as the "gold standard." Self-reported recreational exercise was compared with the following ActiReg-based measures: physical activity energy expenditure (PAEE), minutes of vigorous physical activity (VPA), physical activity level (PAL) and TEE. Pearson's correlations between self-reported weekly exercise and the objectively assessed variables were: rPAEE=0.26, rVPA=0.32, rPAL=0.30 (all P<0.01) and rTEE=0.17 (P=0.07). The partial correlation coefficients between the questionnaire responses and the ActiReg measurements were similar after adjusting for parity, body mass index, education, age, height and smoking, but rTEE increased (r=0.27, P<0.01). We observed significant positive associations between self-reported exercise activities and motion sensor measurements of physical activity, indicating that the questions used for exercise assessment in MoBa may be useful for ranking pregnant women according to the recreational exercise level.

OBJECTIVE

To develop strategies to recruit and retain inactive older adults into a physical activity program.

METHODS

Names of 7378 older adults were obtained from 60 neighborhoods. Then, 6401 potential subjects were matched to telephone numbers and phoned. Subjects meeting the screening criteria were invited to join the program (n = 4209). Walk leaders and social support were used to enhance retention.

RESULTS

Five hundred seventy-three subjects were recruited (260 intervention and 313 control). The respective participation rate was 12.6% (260/2056) and 14.5% (313/2153), with low attrition of 31.9% (83/260) and 24.6% (77/313).

CONCLUSIONS

Effective recruitment and retention strategies were identified.

In the present study, the effect of LPS on different splenic non-lymphoid cells was investigated. Marginal zone (MZ) macrophages, marginal metallophils and interdigitating cells (IDC) were demonstrated using specific monoclonal antibodies in a two-step immunoperoxidase procedure in combination with enzyme histochemistry. The results indicate that the number of marginal zone macrophages decreases markedly after LPS treatment, but is followed by a rapid repopulation as observed by monoclonal antibody staining and selective uptake of FITC-Ficoll. Marginal metallophils are normally located at the inner border of the marginal sinus and can specifically be identified by the monoclonal antibody MOMA-1. Following LPS stimulation, many MOMA-1-positive cells were present in the corona and central parts of the follicles, with decreasing numbers near the marginal sinus. These findings strongly suggest that LPS induces a migration of marginal metallophils towards the follicle centres. Most of the tangible body macrophages in the follicle centres appeared to be slightly MOMA-1-positive, which indicates that marginal metallophils may, at least under certain circumstances, differentiate into tangible body macrophages. In the inner PALS, many interdigitating cells, NLDC-145-positive cells, can be found. The number of NLDC-145-positive cells was shown to be severely decreased at later time-intervals after LPS administration, resulting in an almost unstained inner PALS at 2 days. In contrast to the above-mentioned splenic non-lymphoid cells, the red pulp macrophages are only minimally affected by LPS.

BACKGROUND

Executive function deficits in depression implicate involvement of frontal-striatal circuits. However, studies of hypothalamic-pituitary-axis (HPA) function suggest that stress-related brain changes of hippocampus may also implicate prefrontal-hippocampal circuits, which may explain the profile of both executive dysfunction and memory deficits. In this study we examined the performance of patients with major depressive disorder (MDD) on tasks of memory and executive function in relation to melancholic features and to cortisol levels. Our hypothesis was that raised cortisol levels in melancholic patients would correlate with these deficits.

METHODS

Forty female MDD patients, 20 having melancholic features (MEL vs. Non-MEL), and 20 sex-age- and education-matched normal controls were investigated using the Cambridge neuropsychological test automated battery (CANTAB), to assess memory (paired associative learning, PAL; short-term recognition memory, SRM) and executive (intradimensional/extradimensional set-shifting, ID/ED; Stockings of Cambridge, SOC) functions. Plasma and salivary cortisol levels were measured.

RESULTS

The MDD patients performed worse than controls on PAL and both executive tasks. The MEL group differed from controls on all tests, and differed from the non-MEL only at the ED stage of the ID/ED task. Patient cortisol levels were within the normal range and did not correlate with neuropsychological performance for any group.

CONCLUSIONS

MDD patients showed neuropsychological deficits on tasks of executive function and memory, supporting the model of frontal-temporal dysfunction. MEL vs. non-MEL performed worse overall and demonstrated a qualitative difference in set shifting, perhaps implicating more extensive prefrontal involvement. Cortisol levels did not correlate with depression severity or the observed deficits.

BACKGROUND

Radiographic measurements are often used as a substitute for direct clinical measurements requiring re-entry surgery for follow-up outcome studies.

OBJECTIVE

(1) To assess the reliability of clinical and radiographic measurements of periodontal defects as compared to direct bone measurements during surgical procedures, and (2) to assess the associations between selected clinical and radiographic measurements of periodontal inter-proximal defects.

METHODS

57 inter-proximal periodontal defects were measured at baseline and at 12 months after surgical treatment. Direct measurements during surgery of the distance between the CEJ to the bottom of defects (ABL) were compared with probing to bone (PB), probing attachment level (PAL), and radiographic measurements.

RESULTS

Probing to bone is an accurate measure to assess inter-proximal bone level as compared to ABL (mean difference: 0.1 mm) and that intra-oral standardized radiographs underestimate bone level and defect depth by approximately 1.4 mm. The interpretation of periodontal changes between baseline and 12 months after treatment by probing to bone, or PAL measurements, or from radiographic images yield almost identical results (mean difference< or =0.2 mm). For the assessments of changes over time using PB change as the standard, intra-class correlation (ICC) coefficients varied between 0.52 to 0.90. The best ICC coefficient was found for relative attachment level change assessed by the Florida probe (0.90), and with an ICC value of 0.61 for changes assessed from intra-oral radiographs. Two-way analysis of variance failed to demonstrate differences between sets of comparisons.

CONCLUSIONS

Both radiographic interpretations of changes over time, and measurements of attachment level changes are reliable in assessing the treatment outcome of inter-proximal intra-bony defects when compared to probing to bone changes as the standard method.

Immunological screening of a Pseudomonas aeruginosa cosmid library led to the identification of clones producing an 18 kDa outer-membrane protein. This protein reacted in Western blots with a polyclonal antiserum against outer-membrane proteins of P. aeruginosa and with a monoclonal antibody (MA1-6) specific for OprL, the peptidoglycan-associated outer-membrane lipoprotein (PAL). Sequencing of pOML7, a subclone expressing oprL, revealed an ORF of 504 bp encoding a polypeptide with a typical lipoprotein signal recognition sequence. Another ORF was found upstream of oprL, with homology to the TolB protein of Escherichia coli and Haemophilus influenzae. Downstream of oprL, a second ORF, of 321 bp, was found (orf2), encoding a protein with a signal peptide and with no homology with proteins of known biological function. After the stop codon of orf2, a rho-independent terminator sequence was detected which is part of the P. aeruginosa PAO1 insertion element IS222. OprL showed homologies with all known PALs from Gram-negative bacteria, especially in the C-terminal part. mAb MA1-6 reacted with P. aeruginosa cells in immunofluorescence, and with E. coli cells expressing oprL, which had an abnormal, elongated morphology, an indication that production of the protein perturbed the division process.

This paper examines the catalytic function of the protein YbgC, encoded by the ybgC gene of the tol-pal gene cluster in Haemophilus influenzae. The YbgC protein, a homologue of the Pseudomonas sp. strain CBS3 4-hydroxybenzoyl-coenzyme A thioesterase, conserves the active site Asp residue associated with thioesterase activity. The H. influenzae ybgC gene was cloned and overexpressed in Escherichia coli. The recombinant protein was purified and tested for thioesterase activity towards acyl-CoA and acyl-N-acetylcysteamine thioesters. The YbgC protein catalyzes the hydrolysis of short chain aliphatic acyl-CoA thioesters, while the D18N YbgC mutant protein (prepared to serve as a control) does not.

OBJECTIVE

To measure total energy expenditure (TEE) for normal healthy Japanese by the doubly labelled water (DLW), and to compare the physical activity level (PAL) among categories classified by the categories used in daily reference intake (DRI), Japan and the International Physical Activity Questionnaire (IPAQ).

METHODS

A total of 150 healthy Japanese men and women aged 20- to 59-year-old living in four districts of Japan. TEE was measured by the DLW method, and the PAL was calculated from TEE divided by basal metabolic rate. Simultaneously with TEE measurement, the PAL was assessed employing the categories used in DRI, Japan and IPAQ.

RESULTS

The average TEE and PAL were 10.78+/-1.67 MJ/day and 1.72+/-0.22 for males and 8.37+/-1.30 MJ/day and 1.72+/-0.27 for females, respectively. The subjects in the highly active categories assessed by both DRI and IPAQ showed significantly higher PAL compared with less active categories. However, PALs among light and moderate categories by DRI, and insufficient and sufficiently active by IPAQ were not significantly different.

CONCLUSIONS

In developed countries, highly active subjects could be assessed by a simple questionnaire. However, the questionnaire should be improved to clarify the sedentary to moderately active subjects by assessing carefully very light to moderate physical activity.

OBJECTIVE

We investigated whether an initial neuropsychological assessment could predict rapid progression over 12 months, from amnestic mild cognitive impairment (aMCI) to Alzheimer's disease (AD).

METHODS

A longitudinal study compared the neuropsychological profiles of 27 normal controls and 18 aMCI patients at baseline and 12 months.

RESULTS

At 12 months, 24 control subjects followed up remained cognitively normal. 7 aMCI patients (6 multiple-domain aMCI and 1 single-domain aMCI) progressed to AD, and 11 were non-progressors. Prognosis was best captured by a combination of associative learning, the paired associate learning task (PAL), and global cognition, the Addenbrooke's Cognitive Examination (ACE).

CONCLUSIONS

The PAL and ACE can sensitively detect meaningful differences in scores at baseline and may be used as prognostic indicators. Multiple-domain aMCI patients progressed rapidly to AD and may be more usefully labelled as early stage AD.

OBJECTIVE

The present study was designed to (a) evaluate and compare plasma ammonia levels (PAL) in patients with acute liver failure (ALF) and chronic liver disease (CLD) with or without hepatic encephalopathy (HE); (b) correlate the severity of HE with PAL; and (c) correlate PAL with clinical features of raised intracranial tension in ALF.

METHODS

A total of 40 patients, comprised of 20 patients with ALF (Group A) and 20 patients with CLD (Group B, which was comprised of 8 patients with HE (subgroup B1) and 12 patients without HE (subgroup B2)), were studied. PAL was estimated using an enzymatic UV-method (RANDOX). The clinical and biochemical profile of all the patients was recorded. Correlation between the grade of HE and PAL was derived using Pearson's correlation coefficient. The mean PAL of ALF patients with and without raised intracranial tension was compared using the standard error of difference between the two means.

RESULTS

The mean PAL (micromol/L) +/- SD was as follows: Group A: 172.1 +/- 52.55, subgroup B1: 58.75 +/- 29.38, subgroup B2: 42.17 +/- 18.19 (normal levels = 10-47 micromol/L). All patients with ALF showed PAL more than the upper limit of the normal range, and there was good correlation between the severity of HE and PAL [r = 0.91 at P < 0.05]. In subgroup B1 (CLD with HE), 3/8 patients (37.5%), and in subgroup B2 (CLD with HE), 4/12 patients (33.3%) patients had PAL more than the upper limit of normal range. Within Group A, 14 patients had clinical features of raised intracranial tension/cerebral edema, and the mean PAL of these patients (188.21 +/- 49.15 micromol/L) was significantly higher than those who did not have features of raised intracranial tension (134.5 +/- 42.36 micromol/L) (SE of difference between two means).

CONCLUSIONS

Raised PAL appears to be an important laboratory abnormality seen in patients with ALF, and there seems to be a significant correlation between the severity of encephalopathy and PAL in these patients. However, among patients with CLD, the proportion of patients with PAL more than the upper limit of normal range is not significantly different between those with or without HE. Our study also suggests that high PAL in ALF patients appears to correlate with clinical features of cerebral edema and raised intracranial tension.

Formation of new blood vessels is essential for several physiological and pathological events, e.g. embryogenesis, wound healing and tumor growth and metastasis. In order to increase the insight into the mechanisms of angiogenesis we have visualized the different components of the microvasculature in human wounds and tumors by immunohistochemistry on the light and electronmicroscopic level. For this purpose, antibodies recognizing distinct markers for human endothelial cells, pericytes and basal lamina were used on freshly frozen or paraformaldehyde-fixed tissue samples. In terms of efficacy, the PAL-E antigen is highly specific for blood vessel endothelium. Its sensitivity is less than other endothelial markers, such as von Willebrand factor and CD 31, as it is not expressed in arterioles. Within the context of the microvasculature alpha-smooth muscle actin and the HMW-MAA chondroitin sulphate proteoglycan are useful markers for pericytes. Type IV Collagen and Laminin can be visualized consistently in the microvascular basal lamina. During the formation of granulation tissue in wound healing a heterogeneity of the expression of endothelial and pericyte markers is found. In the least matured zone in granulation tissue of decubitus lesions and experimental skin wounds microvessels already contained both endothelial cells and pericytes, suggesting a role for both cell types in the early steps of angiogenesis. Regarding the tumor microvasculature, antibodies to von Willebrand factor often failed to stain capillaries, that did show expression of the other endothelial markers studied. Broad staining in pericytes was found for the HMW-MAA chondroitin sulphate proteoglycan. In contrast, these cells only locally expressed alpha-smooth muscle actin. Staining of the basal lamina components Type IV Collagen and Laminin within tumors was not restricted to the microvasculature. Therefore, antibodies recognizing endothelial markers, particularly PAL-E and BMA 120, are preferable as tools to visualize the tumor microvasculature. In accordance with the situation in granulation tissue of wound healing the broad presence of pericytes in the microvasculature of human tumor suggests an involvement of this cell type in tumor angiogenesis. Recent immunohistochemical studies on human tumor lesions indicated that a high number of microvessels adjacent to the tumor as a measure of tumor angiogenesis is an unfavorable prognostic factor in cutaneous melanoma, mammary carcinoma and non-small cell pulmonary carcinoma. This new application of immunohistochemistry represents a valuable, clinically relevant adjunct to the repertoire of the surgical pathologist.

BACKGROUND

Persistent air leak (PAL; defined as air leak>> 5 days) after major pulmonary resection is prevalent and associated with significant morbidity. This study examines an incompletely characterized treatment for the management of PAL, chemical pleurodesis.

METHODS

A retrospective case-control study examining all isolated lobectomies and bilobectomies by thoracotomy was performed. The PALs (1997 to 2006) and controls (2002 to 2006) were identified from a prospective database. Incidence, risk factors, management, and outcome were defined.

RESULTS

Over 9 years, 78 PALs were identified in 1,393 patients (5.6%). Controls consisted of 700 consecutive patients. Propensity score analysis matching case and controls showed no predictive risk factors for air leak using a logistic regression model. Univariate analysis demonstrated that female gender, smoking history, and forced vital capacity were predictive risk factors. Treatment of PAL consisted of observation (n = 33, 42.3%), pleurodesis (n = 41, 52.6%), Heimlich valve (n = 3, 3.8%), and reoperation (n = 1, 1.3%). Seventy-three patients (93.6%) required no further intervention. One patient required a muscle flap, one readmission for pneumothorax, and one empyema resulting in death. Sclerosis was successful in 40 of 41 patients (97.6%). Mean time to treatment was 8.4 +/- 3.6 days, mean duration of air leak was 10.7 +/- 4.5, and mean duration of air leak postsclerotherapy was 2.8 +/- 2.2 days. Postoperative pneumonia occurred with increased frequency in PAL patients (6 of 45 [13.3%] vs 34 of 700 [4.9%], p = 0.014). PAL was associated with increased length of stay (14.2 vs 7.1 days, p < 0.001) and time with chest tube (11.5 vs 3.4 days, p < 0.001).

CONCLUSIONS

Air leaks remain an important cause of morbidity. Pleurodesis is an effective option in management of PAL after major pulmonary resection.

A genomic library of Legionella pneumophila, the causative agent of Legionnaires disease in humans, was constructed in Escherichia coli K-12, and the recombinant clones were screened by immuno-colony blots with an antiserum raised against heat-killed L. pneumophila. Twenty-three clones coding for a Legionella-specific protein of 19 kDa were isolated. The 19-kDa protein, which represents an outer membrane protein, was found to be associated with the peptidoglycan layer both in L. pneumophila and in the recombinant E. coli clones. This was shown by electrophoresis and Western immunoblot analysis of bacterial cell membrane fractions with a monospecific polyclonal 19-kDa protein-specific antiserum. The protein was termed peptidoglycan-associated protein of L. pneumophila (Ppl). The corresponding genetic determinant, ppl, was subcloned on a 1.8-kb ClaI fragment. DNA sequence studies revealed that two open reading frames, pplA and pplB, coding for putative proteins of 18.9 and 16.8 kDa, respectively, were located on the ClaI fragment. Exonuclease III digestion studies confirmed that pplA is the gene coding for the peptidoglycan-associated 19-kDa protein of L. pneumophila. The amino acid sequence of PplA exhibits a high degree of homology to the sequences of the Pal lipoproteins of E. coli K-12 and Haemophilus influenzae.