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Rice is one of the most important crops, feeding more than half of the world population. There are two cultivated species, the African rice Oryza glaberrima and the Asian rice O. sativa. Although the African species is gradually replaced by O. sativa in most of African rice agrosystems, this species represents an important reservoir of genes of agronomical interest. Their exploitation for the development of modern African rice varieties requires a good understanding of the genetic relationships between the two cultivated species. We took advantage of the recent availability of the sequence of the chromosome 3 short arm of O. glaberrima to estimate the date of radiation between O. glaberrima and O. sativa lineages, using all the long terminal repeat (LTR)-retrotransposons as paleogenomic markers. We first demonstrated that in two distinct lineages, LTR-retrotransposons mutate at the same rate. Based on LTR-retrotransposons shared by both species in orthologous position, we then estimated that O. glaberrima and O. sativa progenitors diverged 1.2 Ma. This constitutes one of the first studies using such a large sample of transposable elements to reconstruct the phylogeny of species. Given the number of genome sequencing projects, there is no doubt that such approach will allow to resolve phylogenetic incongruities. The application of this method to other plant genomes will also facilitate further understanding of evolution of LTR-retrotransposons and eventually of the whole genome in divergent plant lineages.

We report an improved draft nucleotide sequence of the 2.3-gigabase genome of maize, an important crop plant and model for biological research. Over 32,000 genes were predicted, of which 99.8% were placed on reference chromosomes. Nearly 85% of the genome is composed of hundreds of families of transposable elements, dispersed nonuniformly across the genome. These were responsible for the capture and amplification of numerous gene fragments and affect the composition, sizes, and positions of centromeres. We also report on the correlation of methylation-poor regions with Mu transposon insertions and recombination, and copy number variants with insertions and/or deletions, as well as how uneven gene losses between duplicated regions were involved in returning an ancient allotetraploid to a genetically diploid state. These analyses inform and set the stage for further investigations to improve our understanding of the domestication and agricultural improvements of maize.

Recent comprehensive sequence analysis of the maize genome now permits detailed discovery and description of all transposable elements (TEs) in this complex nuclear environment. Reiteratively optimized structural and homology criteria were used in the computer-assisted search for retroelements, TEs that transpose by reverse transcription of an RNA intermediate, with the final results verified by manual inspection. Retroelements were found to occupy the majority (>75%) of the nuclear genome in maize inbred B73. Unprecedented genetic diversity was discovered in the long terminal repeat (LTR) retrotransposon class of retroelements, with >400 families (>350 newly discovered) contributing >31,000 intact elements. The two other classes of retroelements, SINEs (four families) and LINEs (at least 30 families), were observed to contribute 1,991 and approximately 35,000 copies, respectively, or a combined approximately 1% of the B73 nuclear genome. With regard to fully intact elements, median copy numbers for all retroelement families in maize was 2 because >250 LTR retrotransposon families contained only one or two intact members that could be detected in the B73 draft sequence. The majority, perhaps all, of the investigated retroelement families exhibited non-random dispersal across the maize genome, with LINEs, SINEs, and many low-copy-number LTR retrotransposons exhibiting a bias for accumulation in gene-rich regions. In contrast, most (but not all) medium- and high-copy-number LTR retrotransposons were found to preferentially accumulate in gene-poor regions like pericentromeric heterochromatin, while a few high-copy-number families exhibited the opposite bias. Regions of the genome with the highest LTR retrotransposon density contained the lowest LTR retrotransposon diversity. These results indicate that the maize genome provides a great number of different niches for the survival and procreation of a great variety of retroelements that have evolved to differentially occupy and exploit this genomic diversity., Competing Interests: The authors have declared that no competing interests exist.

Most of our understanding of plant genome structure and evolution has come from the careful annotation of small (e.g., 100 kb) sequenced genomic regions or from automated annotation of complete genome sequences. Here, we sequenced and carefully annotated a contiguous 22 Mb region of maize chromosome 4 using an improved pseudomolecule for annotation. The sequence segment was comprehensively ordered, oriented, and confirmed using the maize optical map. Nearly 84% of the sequence is composed of transposable elements (TEs) that are mostly nested within each other, of which most families are low-copy. We identified 544 gene models using multiple levels of evidence, as well as five miRNA genes. Gene fragments, many captured by TEs, are prevalent within this region. Elimination of gene redundancy from a tetraploid maize ancestor that originated a few million years ago is responsible in this region for most disruptions of synteny with sorghum and rice. Consistent with other sub-genomic analyses in maize, small RNA mapping showed that many small RNAs match TEs and that most TEs match small RNAs. These results, performed on approximately 1% of the maize genome, demonstrate the feasibility of refining the B73 RefGen_v1 genome assembly by incorporating optical map, high-resolution genetic map, and comparative genomic data sets. Such improvements, along with those of gene and repeat annotation, will serve to promote future functional genomic and phylogenomic research in maize and other grasses., Competing Interests: The authors have declared that no competing interests exist.

Transposable elements are ubiquitous components of plant genomes. When active, these mobile elements can induce changes in the genome at both the structural and functional levels. Availability of the complete genome sequence for several model plant species provides the opportunity to study TEs in plants at an unprecedented scale. In the case of rice, annotation of the genomic sequence of the variety Nipponbare has revealed that TE-related sequences form more than 25% of its genome. However, most of the elements found are inactive, either because of structural alterations or because they are the target of various silencing pathways. In this paper, we propose a new post-genomic strategy aimed at identifying active TEs. Our approach relies on transcript profiling of TE-related sequences using a tiling microarray. We applied it to a particular class of TEs, the LTR retrotransposons. A transcript profiling assay of rice calli led to identification of a new transpositionally active family, named Lullaby. We provide a complete structural description of this element. We also show that it has recently been active in planta in rice, and discuss its phylogenetic relationships with Tos17, the only other active LTR retrotransposon described so far in the species.

The passenger lymphocyte syndrome (PLS) is an unusual complication of solid organ transplantation, in which donor lymphocytes produce antibodies reactive with host red blood cell (RBC) antigens. Risks for PLS include highly lymphoid grafts, past sensitization of the donor against relevant RBC antigens, and donor lymphocyte escape of host immune clearance. For a 1-year period at our center, we observed an uncommonly high frequency of post-lung transplant Rhesus PLS, occurring once in every 31 cases. Passenger lymphocyte syndrome resulted from 2 alloimmunized cadaveric donors, in 3 of 3 D+, ABO-identical but HLA-unmatched recipients who initially had nonreactive RBC antibody screens. In case 1, the right lung of a group A, D-negative donor, with antibodies against D, C, and E antigens, was transplanted into a group A, R1R1 recipient. The recipient developed severe hemolytic anemia, direct antiglobulin test (DAT)-positive, on postoperative day (POD) 17. Anti-D and anti-C were identified on both the indirect antiglobulin test (IAT) and the RBC eluate. She required 10 U of RBCs in 40 days as well as plasmapheresis (POD 36-40). When transfusion dependence ceased, anti-D +/- C remained detectable on DAT and IAT for another 6.5 months. In case 2, the group A, R1r recipient of the same donor's left lung exhibited anti-D for the first time at posttransplant month 4 on both IAT and DAT. This activity persisted until a rejection episode 5 months later, without ever causing any evidence of hemolysis. In case 3, the group O, R1R1 recipient of both lungs of a group O, D-negative donor, with antibodies against D, C, and V antigens, developed a nonhemolytic DAT and IAT with anti-D +/- C at postoperative month 2, which remained positive at last follow-up (6 months posttransplant). In conclusion, this report suggests a high incidence of Rhesus antibody PLS after lung transplantation, with wide variations in the timing of antibody onset, persistence, and severity. A review of the phenomenon and its implications are discussed.

Background: Efforts to develop global programs for the control of iron deficiency require simple, low-cost, and accurate indicators of iron status., Objective: We aimed to compare estimates of body iron (BI) stores, as calculated from either plasma ferritin concentration alone (BI-ferritin) or the ratio of plasma transferrin receptor (TfR) to ferritin (BI-TfR/ferritin)., Design: Data were analyzed from 4 previously completed, randomized intervention trials that enrolled infants, schoolchildren, or pregnant women (total n = 1189, after excluding subjects with elevated C-reactive protein)., Results: The correlation coefficients between BI-ferritin and BI-TfR/ferritin were >0.95 for all studies. The kappa index ranged from 0.5 to 1.0. All of the sensitivities of BI-ferritin for identifying persons with low iron stores (defined as BI-TfR/ferritin < 0 mg/kg body wt) were >0.90. All of the specificities were >0.90 except the study of pregnant women (specificity = 0.66). The effect sizes of iron intervention trials were significantly greater for change in iron reserves estimated by BI-TfR/ferritin than by BI-ferritin in 2 studies with larger effect sizes (1.11 compared with 1.00 and 1.56 compared with 1.44, respectively; P < 0.05) and 1 study with medium effect size (0.70 compared with 0.57; P < 0.05). However, there were no significant differences between estimates of these effect sizes for 1 study with a medium effect size and 1 study with a smaller effect size (0.78 compared with 0.83 and 0.37 compared with 0.35, respectively; P > 0.2)., Conclusion: Plasma ferritin concentration alone provides a good approximation of total BI reserves, as estimated by BI-TfR/ferritin, on the basis of high correlation, sensitivity, and specificity among nonpregnant persons with unelevated C-reactive protein.

The Rice Annotation Project Database (RAP-DB) was created to provide the genome sequence assembly of the International Rice Genome Sequencing Project (IRGSP), manually curated annotation of the sequence, and other genomics information that could be useful for comprehensive understanding of the rice biology. Since the last publication of the RAP-DB, the IRGSP genome has been revised and reassembled. In addition, a large number of rice-expressed sequence tags have been released, and functional genomics resources have been produced worldwide. Thus, we have thoroughly updated our genome annotation by manual curation of all the functional descriptions of rice genes. The latest version of the RAP-DB contains a variety of annotation data as follows: clone positions, structures and functions of 31 439 genes validated by cDNAs, RNA genes detected by massively parallel signature sequencing (MPSS) technology and sequence similarity, flanking sequences of mutant lines, transposable elements, etc. Other annotation data such as Gnomon can be displayed along with those of RAP for comparison. We have also developed a new keyword search system to allow the user to access useful information. The RAP-DB is available at: http://rapdb.dna.affrc.go.jp/ and http://rapdb.lab.nig.ac.jp/.

Background: The impact of panresistant bacteria, other than Burkholderia cepacia, on the survival after lung transplantation in patients with cystic fibrosis (CF) remains controversial., Methods: To determine the impact of panresistant bacteria in CF patients on survival after lung transplantation a retrospective multicenter study was performed. All lung transplant recipients with a pre-transplant diagnosis of CF, at the University of Toronto (n = 53) and Duke University (n = 50), were included. Patients were included in the panresistant group if at least one specimen isolated from their respiratory secretions grew bacteria resistant or intermediate to all classes of antibiotics tested. Patients with sensitive or resistant B cepacia were excluded because of its adverse impact upon post-transplant survival., Results: Forty-five of 103 (43.7%) patients harbored panresistant bacteria (43 had Pseudomonas aeruginosa, 1 had Stenotrophomonas maltophilia, and 1 had Achromobacter xylosoxidans). According to log-rank test, there was decreased survival in patients with panresistant bacteria compared to patients with sensitive bacteria (survival: 91.1 +/- 4.2% vs 98.3 +/- 1.7% at 3 months; 88.6 +/- 4.8% vs 96.6 +/- 2.4% at 1 year; 63.2 +/- 8.6% vs 90.7 +/- 4.0% at 3 years; 58.3 +/- 9.2% vs 85.6 +/- 5.2% at 5 years; p = 0.016). The results did not differ significantly between the two centers. Both groups had similar or better survival than CF patients as reported by the United Network of Organ Sharing (UNOS) registry (1-year, 86.0%; 3 years, 65.4%; 5 years, 49.6%)., Conclusions: Patients with CF harboring panresistant bacteria have slightly decreased survival, but their survival is comparable to the results published by the UNOS registry.

Selection criteria for organ transplantation have evolved over time. Age has been revisited periodically. We studied the outcome of lung transplant adjusted by age in a single center transplant population. We matched the 42 lung graft recipients older than 60 years transplanted by July 1999 to younger controls by lung disease, transplant era within 2 years, type of transplant and gender. The female to male ratios were 17/25 among the older cohort (median age 61.6 years), and 15/27 (median age 51.9 years) among the matched younger. Survival analysis demonstrated a significant difference: at 1 year, 60% versus 86%, and at 5 years, 37% versus 57%, for older and younger, respectively, p=0.005. Excess annual mortality, calculated with the declining exponential approximation to life expectancy (DEALE), showed an older/younger ratio of 1.9. Eleven deaths occurred within 6 months among the older patients, 10 due to infection. After 6 months, there were 20 more deaths, 6 due to malignancy, 5 to Bronchiolitis Obliterans Syndrome (BOS), 3 to infection and 6 to other causes. Among the younger there were 6 deaths within 6 months and 12 more thereafter; among the latter, 8 were due to BOS. Despite stringent selection, lung transplant recipients older than 60 years show increased mortality even after adjusting for their expected higher age-related mortality.

Objective: We examined the outcome of lung transplantation with the use of donors aged 60 years or more., Methods: From May 1994 to May 2005, 467 lung transplants were performed at our institution. A total of 60 recipients received lungs from donors aged 60 years or more (range 60-77 years, median 65 years), whereas 407 recipients received lungs from younger donors (range 9-59, median 39 years)., Results: A total of 48 patients (10%) died within 30 days of surgery: 10 (17%) in the older donor group versus 38 (9%) in the younger donor group (P = .08). The operative mortality varied with the underlying lung disease and was higher in recipients presenting with pulmonary hypertension and pulmonary fibrosis than with emphysema or cystic fibrosis. A total of 210 patients died after a median follow-up of 25 months (range 0-136 months). The overall 5- and 10-year survivals were 57% and 38%, respectively. However, the 10-year survival tended to be worse in the older donor group (16% vs 39% in the younger donor group, P = .07). Bronchiolitis obliterans syndrome was the predominant cause of death in recipients of older donors who survived for more than 90 days after surgery (11/17, 65% vs 45/132, 34% in recipients of younger donors surviving for >90 days after surgery, P = .01)., Conclusions: Given the lack of organ donors, lungs from donors aged 60 years or more should be considered for transplantation. However, the use of donors aged 60 years or more is associated with a lower 10-year survival, and bronchiolitis obliterans syndrome plays a significant role as the cause of late death.

Monoamine oxidase (MAO) is a flavin adenine dinucleotide (FAD)-containing enzyme located at the outer membranes of mitochondria that catalyzes the oxidative deamination of biogenic and xenobiotic amines. We have used a chromatographic method to measure MAO-enzymatic activity by using kynuramine as a non-selective substrate with its MAO-oxidation product subsequently analyzed by RP-HPLC-DAD and HPLC-mass spectrometry (MS). This method was applied to study the kinetic parameters, inhibition and reaction products of MAO recombinant enzymes in presence of tetrahydro-beta-carboline and beta-carboline alkaloids occurring in foods, plants and mammals. Analysis by HPLC showed that tetrahydro-beta-carbolines or beta-carbolines were not modified by MAO. Several beta-carbolines such as tryptoline (1,2,3,4-tetrahydro-beta-carboline) and 1-methyltryptoline (1-methyl-1,2,3,4-tetrahydro-beta-carboline) were inhibitors of MAO-A; instead their corresponding 6-hydroxy-derivatives (6-hydroxytryptoline and 6-hydroxy-1-methyltryptoline) lacked this activity. Tetrahydro-beta-carboline-3-carboxylic acids were unable to inhibit MAO enzymes. In contrast, their oxidation products, i.e. the fully aromatic beta-carbolines (norharman and harman), acted as good inhibitors of MAO. Two tetrahydro-beta-carbolines (i.e. tryptoline and 1-methyltryptoline) occurring in foods were isolated by solid-phase extraction (SPE) and RP-HPLC from selected samples of sausages and the corresponding extracts exhibited good inhibition properties over MAO-A. These results suggest that beta-carbolines from foods, plants, and mammals may exert inhibitory actions on MAO enzymes.

While current donor selection with clinical findings is generally effective, the imprecise nature of the assessment forces clinicians to remain on the conservative side. A reliable biological marker would assist donor selection and would improve donor organ utilization. We collected biopsies from 169 donor lungs before implantation. Expression levels of IL-6, IL-8, IL-10, TNF-alpha, IFN-gamma and IL-1beta were measured by quantitative real-time RT-PCR (qRT-PCR). Seventeen cases died within 30 days after transplantation. No donor factor was significantly associated with 30-day mortality. Univariate analysis of the 84 cases for development of the prediction model showed that IL-6, IL-8, TNF-alpha and IL-1beta were risk factors for mortality and IL-10 and IFN-gamma were protective factors. We analyzed the cytokine expression ratios of risk to protective cytokines. A stepwise logistic regression for 30-day mortality demonstrated that a model containing the ratio of IL-6/IL-10 was the most predictive (p = 0.0013). When applied to the remaining 85 cases for validation, the test of model fit was significant (p = 0.039). Using the cytokine ratio, we were able to define three risk groups with striking differences in survival (p = 0.0003). Multi-cytokine analysis of the donor lung graft with qRT-PCR shows significant promise as a strategy to biologically evaluate the donor lung prior to implantation.

Monoamine oxidase (MAO) is a mitochondrial outer-membrane flavoenzyme involved in brain and peripheral oxidative catabolism of neurotransmitters and xenobiotic amines, including neurotoxic amines, and a well-known target for antidepressant and neuroprotective drugs. Recent epidemiological studies have consistently shown that coffee drinkers have an apparently lower incidence of Parkinson's disease (PD), suggesting that coffee might somehow act as a purported neuroprotectant. In this paper, "ready to drink" coffee brews exhibited inhibitory properties on recombinant human MAO A and B isozymes catalyzing the oxidative deamination of kynuramine, suggesting that coffee contains compounds acting as MAO inhibitors. MAO inhibition was reversible and competitive for MAO A and MAO B. Subsequently, the pyrido-indole (beta-carboline) alkaloids, norharman and harman, were identified and isolated from MAO-inhibiting coffee, and were good inhibitors on MAO A (harman and norharman) and MAO B (norharman) isozymes. beta-carbolines isolated from ready-to-drink coffee were competitive and reversible inhibitors and appeared up to 210 microg/L, confirming that coffee is the most important exogenous source of these alkaloids in addition to cigarette smoking. Inhibition of MAO enzymes by coffee and the presence of MAO inhibitors that are also neuroactive, such as beta-carbolines and eventually others, might play a role in the neuroactive actions including a purported neuroprotection associated with coffee consumption.

Previous studies suggest that bilateral (BLT) compared with single lung transplantation (SLT) for patients with chronic obstructive pulmonary disease (COPD) results in improved long-term survival. The effect of transplant operation on bronchiolitis obliterans syndrome (BOS) is unknown. A retrospective study of all lung transplant recipients with pre-transplant diagnoses of COPD at the University of Toronto and at Duke University was performed. Data collected were age, gender, date and type of transplant, acute rejection, survival, presence and time of BOS. 221 (bilateral n = 101, single n = 120) patients met our criteria. Patients with BLT were younger (53.0 vs. 55.3 years; p = 0.034), more likely to be male (56.3% vs. 42.4%; p = 0.039) and more likely to be transplanted at the University of Toronto (79.6% vs. 16.1%; p < 0.001). Freedom from BOS was similar at 1 year post-transplant. However, BLT recipients were more commonly free from BOS 3 years (57.4% vs. 50.7%) and 5 years (44.5% vs. 17.9%) post-transplant (p = 0.024). Survival of BLT was better than SLT recipients at 3 and 5 years post-transplant (BLT vs. SLT: 67.5% vs. 61.1% and 60.7% vs. 34.1%, respectively; p = 0.018). Similar trends on survival were observed after development of BOS. BLT results in lower rates of BOS in patients with COPD that are eligible for both SLT and BLT.

Cystic fibrosis (CF) related diabetes mellitus (DM) occurs in 15% of adult pancreatic insufficient CF patients. Lung transplantation is a treatment option for end-stage CF. We hypothesized that the prevalence of DM increases after lung transplantation. The study population included adult patients undergoing lung transplantation from March 1988 to March 2002 for end-stage CF at the University of Toronto. Demographic data, exocrine pancreatic function, presence of DM before and after transplant, as well as timing of its development after transplant were collected. Eighty-six patients met the study criteria; 77 of 86 (89.5%) of patients were pancreatic insufficient and were further analyzed. Median follow-up post-transplant was 3.3 yr (interquartile range: 1.2-7.2). Their mean age was 29.7 +/- 8.1 yr and 46 of 77 (59.7%) were male. The prevalence of DM increased from 22 of 77 (28.6%) before transplant to 38 of 77 (49.4%) after transplant (p = 0.008). The median time of DM development after transplant was 80 d (range: 13-4352). Sixteen of 55 (29.1%) of pancreatic insufficient patients who were non-diabetic prior to transplant, developed DM after transplant. DM is common in CF patients undergoing lung transplantation and the prevalence increases after transplant.

Background: Lung transplantation is an established treatment modality for a number of chronic lung diseases. Long-term survival after lung transplantation is limited by chronic allograft dysfunction, usually manifested by bronchiolitis obliterans syndrome. We describe a case series with upper lobe fibrosis, a novel presentation of chronic allograft dysfunction., Methods: We reviewed lung transplants at the Toronto General Hospital and Duke University Hospital from 1990 to 2002 and identified patients with upper lobe fibrosis., Results: Thirteen of 686 patients (6 women) developed upper lobe fibrosis (Toronto, 9; Duke, 4); 12 of 13 had bilateral transplants. The median age at diagnosis was 42 years (range, 19-70). Primary diagnoses were cystic fibrosis, 6; emphysema, 4; sarcoidosis, 1; and pulmonary fibrosis, 2 patients. Radiographic diagnosis was made at a median of 700 days post-transplant (range, 150-2,920). Pulmonary function tests demonstrated predominantly a progressively worsening restrictive pattern. Open lung biopsy specimens revealed dense interstitial fibrosis, with occasional features of obliterative bronchitis, bronchiolitis obliterans obstructive pneumonia, and aspiration. Nine patients died at a median follow-up of 2,310 days (range, 266-3,740), 8 due to respiratory failure., Conclusion: Upper lobe fibrosis is a novel presentation of chronic allograft dysfunction in lung transplant recipients and is differentiated from bronchiolitis obliterans syndrome on the basis of physiologic and radiologic findings.

Background: The presence of antibodies to human leukocyte antigens (HLA) prior to transplantation has been linked to worse post-transplant outcomes in many solid organ transplants. The effect of these antibodies is less clear in lung transplant recipients, although previous studies have suggested an increased incidence of allograft dysfunction., Methods: A retrospective study of all first lung transplant recipients from the University of Toronto (November 1983-July 2001, n = 380) and Duke University (April 1992-June 2000, n = 276) was performed. Demographic data, survival information, and level of last pre-transplant panel reactive antibody (PRA) were collected. PRA level was measured by the complement-dependent cell cytotoxicity assay at both centers. Survival analysis was performed using the Kaplan-Meier method, and groups were compared with the Wilcoxon rank sum test., Results: Of 656 lung transplant recipients, 101 (15.4%) had a PRA greater than 0, 37 (5.6%) had a PRA greater than 10%, and 20 (3.0%) had a PRA greater than 25%. Patients with a PRA greater than 25% had decreased median survival than did the rest of the patients (1.5 vs 5.2 years) and at 1 month (70% vs 90%), 1 year (65% vs 76%), and 5 years (31% vs 50%), respectively (p = 0.006, Wilcoxon's rank sum test) test)., Conclusion: Significant elevation of PRA prior to lung transplantation is associated with worse survival, especially in the early post-transplant period. This may be due to a direct effect of anti-HLA antibodies on the allograft. The effectiveness of treatments such as plasmapheresis and intravenous immunoglobulin prior to transplantation needs to be evaluated.

Background: Aspiration of gastroesophageal refluxate may contribute to lung transplant bronchiolitis obliterans syndrome (BOS). We investigated bile acids in bronchoalveolar lavage fluid (BALF) and studied its role in BOS., Materials and Methods: Surveillance pulmonary function tests and BALF were evaluated in 120 lung recipients. BOS-(0p-3) was diagnosed after 6 months' survival. BOS was defined as "early" if diagnosed within 12 months after a transplant. BALF was assayed for differential cell count, bile acids, and interleukins 8 and 15. Bile acids were considered elevated if greater than normal serum levels ( or =8 micromol/L)., Results: Elevated BALF bile acids were measured in 20 (17%) of 120 patients. BOS was diagnosed in 36 (34%) of 107 patients and judged "early" in 21 (57%) of 36. Median BALF bile acid values were 1.6 micromol/L (range, 0-32 micromol/L) in BOS patients and 0.3 micromol/L (range, 0-16 micromol/L) in non-BOS patients ( P = .002); 2.6 micromol/L (range, 0-32 micromol/L) in early BOS patients and 0.8 micromol/L (range, 0-4.6 micromol/L) in late BOS patients, ( P = .02). Bile acids correlated with BALF IL-8 and alveolar neutrophilia (r = 0.3, P = .0004, and r = 0.3, P = .004, respectively), but not with IL-15. Freedom from BOS was significantly shortened in patients with elevated BALF bile acids (Cox-Mantel test, P = .0001)., Conclusions: Aspiration of duodenogastroesophageal refluxate is prevalent after lung transplantation and is associated with the development of BOS. Elevated BALF bile acids may promote early BOS development via an inflammatory process, possibly mediated by IL-8 and alveolar neutrophilia.

Most patients with cystic fibrosis (CF) and pancreatic insufficiency require supplementation with fat-soluble vitamins to maintain normal serum levels. Even with supplementation, toxicity is rare. We evaluated serum vitamin A and E levels in 23 adult patients with CF who underwent double lung transplantation. Twenty-one of the subjects were pancreatic insufficient. Fifteen subjects had serum vitamin levels before and after transplant. The median time posttransplantation for these subjects was 9 months. Mean serum vitamin A and E levels were significantly higher posttransplantation (P<0.0001, P<0.001, respectively). Eight subjects who only had posttransplant vitamin levels also had abnormally high vitamin A levels. Although the etiology of this novel finding is unclear, possibilities include altered absorption, drug interactions, impaired retinol metabolism, or increased hepatic synthesis of retinol binding protein.

Monoamine oxidase (MAO) is a mitochondrial outer-membrane flavoenzyme involved in brain and peripheral oxidative catabolism of neurotransmitters and xenobiotic amines, including neurotoxic amines, and a well-known target for antidepressant and neuroprotective drugs. Recently, positron emission tomography imaging has shown that smokers have a much lower activity of peripheral and brain MAO-A (30%) and -B (40%) isozymes compared to non-smokers. This MAO inhibition results from a pharmacological effect of smoke, but little is known about its mechanism. Working with mainstream smoke collected from commercial cigarettes we confirmed that cigarette smoke is a potent inhibitor of human MAO-A and -B isozymes. MAO inhibition was partly reversible, competitive for MAO-A, and a mixed-type inhibition for MAO-B. Two beta-carboline alkaloids, norharman (beta-carboline) and harman (1-methyl-beta-carboline), were identified by GC-MS, quantified, and isolated from the mainstream smoke by solid phase extraction and HPLC. Kinetics analysis revealed that beta-carbolines from cigarette smoke were competitive, reversible, and potent inhibitors of MAO enzymes. Norharman was an inhibitor of MAO-A (K(i)=1.2+/-0.18 microM) and MAO-B (K(i)=1.12+/-0.19 microM), and harman of MAO-A (K(i)=55.54+/-5.3nM). Beta-carboline alkaloids are psychopharmacologically active compounds that may occur endogenously in human tissues, including the brain. These results suggest that beta-carboline alkaloids from cigarette smoke acting as potent reversible inhibitors of MAO enzymes may contribute to the MAO-reduced activity produced by tobacco smoke in smokers. The presence of MAO inhibitors in smoke like beta-carbolines and others may help us to understand some of the purported neuropharmacological effects associated with smoking.

Background: Lung infection with Burkholderia cepacia complex before lung transplantation in patients with cystic fibrosis is a major risk factor for decreased post-operative survival rates compared with those of patients colonized with the more common opportunistic pathogen Pseudomonas aeruginosa. Because adherence to mucosal surfaces is an important initial step in infection, we investigated the use of non-toxic neutral polysaccharides and a sugar alcohol to prevent adherence of B cepacia complex to allograft airway epithelium., Methods: We used human airway explants prepared from donor tracheobronchial tissue to test the effect of dextrans and xylitol in inhibiting the binding of Burkholderia cepacia complex. We used immunofluorescence and electron microscopy to determine the distribution of bacteria in the explants., Results: Burkholderia cepacia complex bound to the explants and was found only in the surface mucus layer. Dextran 40 kd applied before adding the bacteria decreased the number of bound organisms by 80% to 99%. Smaller molecular mass dextrans (4 and 20 kd) were ineffective. Xylitol inhibited bacterial binding by 67% to 85%. Both agents seemed to decrease the thickness of the surface mucus, suggesting that they may indirectly inhibit bacterial binding by removing adherent surface mucus., Conclusions: Treating donor lungs with dextran 40 kd or xylitol before (and possibly after) surgery may inhibit the adherence of Burkholderia cepacia complex to airways and may prevent or decrease subsequent infection of the allografts.

Objectives: The objective of this study was to examine the long-term patient outcomes of lung transplantation in a single center., Methods: Between 1983 and 2003, 521 lung transplants were performed in 501 patients. Major indications were cystic fibrosis (n = 124), chronic obstructive pulmonary disease (n = 88), alpha-1 antitrypsin deficiency (n = 63), pulmonary fibrosis (n = 97), primary pulmonary hypertension (n = 35), Eisenmenger syndrome (n = 21), and miscellaneous end-stage lung diseases (n = 93)., Results: The 5-, 10-, and 15-year survivals for all recipients were 55.1% (95% confidence interval: +/-5%), 35.3% (+/-6%), and 26.5% (+/-11%), respectively. The most common causes of death were sepsis and bronchiolitis obliterans syndrome. Despite an increased postoperative mortality rate, patients with primary pulmonary hypertension achieved the best long-term survival (10-year survival: 59%). Recipients with cystic fibrosis without Burkholderia cepacia infection achieved significantly better long-term survival (10-year survival: 52%) than those with Burkholderia cepacia infection (10-year survival: 15%). The 10-year survival was also significantly better in recipients with chronic obstructive pulmonary disease (43%) than in recipients with alpha-1 antitrypsin deficiency (23%). Although the incidence of bronchiolitis obliterans syndrome was similar between recipients with chronic obstructive pulmonary disease (39%) and alpha-1 antitrypsin deficiency (46%), recipients with alpha-1 antitrypsin deficiency died of sepsis more frequently than recipients with chronic obstructive pulmonary disease (27% vs 6%, respectively; P =.0003)., Conclusions: Although bronchiolitis obliterans syndrome and sepsis still limit the durability of the benefit, lung transplantation returns many patients with end-stage lung disease to active and productive lives. Differences in the complications and long-term survival show the important contribution of the recipient diagnosis to the success of lung transplantation.

Purpose: To determine whether there are thin-section computed tomographic (CT) features that predict bronchiolitis obliterans syndrome (BOS) in lung transplant recipients before the clinical appearance and during the early stages of the disease., Materials and Methods: Two hundred ninety-eight thin-section CT scans obtained in 26 lung transplant recipients who did (study group) and 26 lung transplant recipients who did not (control group) develop BOS were reviewed for the presence of mosaic perfusion, bronchiectasis, bronchial wall thickening, and air trapping. BOS was defined by using the recently revised definition of the International Society for Heart and Lung Transplantation. CT scans obtained in the BOS group were divided into three groups: Group A consisted of the last scans obtained before the clinical appearance of BOS; groups B and C consisted of, respectively, the first and last scans obtained after the clinical appearance of BOS. Scans obtained in the control group were acquired during similar posttransplantation periods and matched to scans in each BOS group. Sensitivity, specificity, and positive and negative predictive values were calculated separately for each subgroup. The optimal threshold for each thin-section CT-depicted abnormality was defined by using receiver operating characteristics analysis., Results: The sensitivities of air trapping for the diagnosis of BOS during the periods in which the scans in groups A, B, and C were obtained were 50%, 44%, and 64%, respectively; specificities were 80%, 100%, and 80% respectively. Sensitivities of mosaic perfusion were 4%, 20%, and 36%, respectively; specificities were 100%, 96%, and 96%, respectively. Sensitivities of bronchiectasis were 25%, 24%, and 32%, respectively; specificities were 80%, 80%, and 96%, respectively. Sensitivities of bronchial wall thickening were 4%, 24%, and 40%, respectively; specificities were 96%, 84%, and 80%, respectively. Air trapping was seen intermittently in nine (43%) of 21 patients with CT scans that depicted this finding at least once., Conclusion: The value of the finding of air trapping before the clinical appearance and during the early stages of BOS is lower than has been previously reported. When using the recently revised criteria for BOS, the role of thin-section CT as a screening test to evaluate patients with lung transplants appears to be limited.

Objective: A retrospective study involving 13 institutions was performed to assess the efficacy of conversion from cyclosporine (INN: ciclosporin) to tacrolimus., Methods: Data from 244 patients were analyzed. Indications for conversion were recurrent-ongoing rejection (n = 110) and stage 1 to 3 bronchiolitis obliterans syndrome (n = 134)., Results: The incidence of acute rejection decreased significantly within 3 months after versus before the switch from cyclosporine to tacrolimus (P <.01). For patients with recurrent-ongoing rejection, the forced expiratory volume in 1 second decreased by 1.96% of predicted value per month (P =.08 vs zero slope) before and increased by 0.34% of predicted value per month (P =.32 vs zero slope) after conversion (P <.06). For patients with stage 1 to 3 bronchiolitis obliterans syndrome, a significant reduction of rejection episodes was observed (P <.01). In single transplant recipients a decrease of the forced expiratory volume in 1 second averaged 2.25% of predicted value per month (P <.01 vs zero slope) before and 0.29% of predicted value per month after conversion. Corresponding values for bilateral transplant recipients were 3.7% of predicted value per month (P <.01 vs zero slope) and 0.9% of predicted value per month (P = 0.04 vs zero slope), respectively. No significant difference in the incidence of infections within 3 months before and after conversion was observed., Conclusions: Conversion from cyclosporine to tacrolimus after lung transplantation is associated with reversal of recurrent-ongoing rejection. Conversion for bronchiolitis obliterans syndrome allows short-term stabilization of lung function in most patients.

Numerous sites have been polluted with mercury as a result of accidental spills and improper disposal practices, and these mercury-contaminated sites may have adverse effects on human health and the environment. Innovative and cost-effective remediation techniques are urgently needed, and this study was performed to investigate the use of electrokinetics for mercury-contaminated soils. Initially, batch tests were performed on two soils, kaolin and glacial till, spiked with mercury(II) to investigate mercury desorption and complexation under different pH environments (pH range 2-12). The complexing agents included disodium ethylenediaminetetraacetate (Na-EDTA), potassium iodide (KI), and sodium chloride (NaCl), and these solutions were used at a concentration of 0.1 M. In addition, deionized water was used for comparison purposes. Based on the batch tests, Na-EDTA and KI were identified as the complexing agents with the greatest potential. The removal efficiency of these complexing agents was then examined by conducting electrokinetic experiments that employed the same solution concentration (0.1 M) and voltage gradient (1.0 VDC/cm) conditions. These tests indicated that for both soils, KI was a more effective complexing agent than Na-EDTA under electrokinetics. For the kaolin soil, the electrokinetic treatment using KI removed approximately 97% of the initial contaminant present (500mg/kg of Hg(ll)), leaving a residual concentration of 16 mg/kg of Hg in the soil, whereas on the glacial till soil, KI removed only 56% of the initial contaminant present (500mg/kg of Hg(II)), leaving a residual concentration of 220 mg/kg of Hg in the soil. The lower Hg removal from glacial till is attributed to the presence of organic matter, which increased mercury adsorption or the formation of insoluble mercury surface complexes.

Study Objective: Lung transplantation is now available for patients with cystic fibrosis (CF) and end-stage lung disease. While pulmonary graft function is often considered the major priority following transplantation, the nonpulmonary complications of this systemic disease also continue. We examined the GI complications in a cohort of patients who underwent transplantation., Design: This was a retrospective study of all patients with CF who underwent transplantation between March 1988 and December 1998 in Toronto. Medical records were reviewed, and a short questionnaire was mailed to patients who were alive as of December 1998., Results: There were 80 bilateral lung transplants performed in 75 patients. The questionnaire was distributed to 43 patients, of whom 27 patients (63%) responded. Pancreatic insufficiency requiring enzyme intake was evident in 72 of 75 patients (96%) at the time of surgery. Of three pancreatic-sufficient patients (4%), pancreatic insufficiency was diagnosed in two patients later. Biliary cirrhosis was diagnosed in three patients prior to transplantation. Distal intestinal obstruction syndrome (DIOS) was recorded for 15 patients (20%). Ten patients had a single episode, of which eight episodes occurred early in the postoperative period. Five patients had recurrent episodes. All were medically treated, except for two patients who underwent surgery. Other complications included cholecystitis (n = 3), mucocele of the appendix (n = 1), peptic ulcer disease (n = 3), and colonic carcinoma (n = 1)., Conclusion: GI complications after lung transplantation are common in patients with CF, and attention should be paid to the risk for DIOS in the early postoperative period. Prevention and early medical treatment are important in order to avoid acute surgery. Close collaboration with the CF clinic, in order to diagnose and treat CF-related complications, is recommended.

Background: Posttransplant lymphoproliferative disease (PTLD) is now a widely recognized complication of lung transplantation. In the current study, we present our experience with PTLD over a 15-year period, which includes the incidence rates in 242 lung allografts and the relative risk of developing PTLD in 146 patients with known pretransplantation Epstein-Barr virus (EBV) status., Methods: Inpatient and outpatient charts of 300 consecutive lung transplant recipients between 1984 and 1999 were retrospectively reviewed., Results: Twelve cases of PTLD were observed for a total incidence rate of 5.0%. Ten of these patients had pretransplantation EBV testing, and the consequent increase in relative risk for patients who were EBV negative was 6.8-fold. The mean time between organ transplantation and tissue diagnosis of PTLD was 17.6 months. Total 1-year survival rate from the time of diagnosis for the cohort was 58%, whereas 2-year survival rate was 50%. Median survival for the six patients who died was 4.5 months., Conclusions: These data suggest that although EBV seronegativity does carry a 6.8-fold increase in the relative risk of developing PTLD, long-term survival despite the development of PTLD can be achieved, and thus EBV seronegativity by itself should not be considered a contraindication to lung transplantation.

The fibrolipoma is a benign tumor variant of the lipoma, characterized by the presence of adipose and fibrous tissues. The authors report a case of a big oral fibrolipoma in a 72 year old woman. After surgery, a mass of 13 x 8 x 6 cm was obtained. The tumor had an implantation pedicle of 1 cm, on the floor of the mouth. The microscopic evaluation showed the presence of polygonal cells grouped into nests and separated by fibrous connective tissue septa. We have not found any report in the literature related to a fibrolipoma located on the floor of the mouth with the characteristics presented in this work.

As a result of concern over excessive mortality after lung transplantation, many transplant programs refuse to accept cystic fibrosis (CF) patients infected with Burkholderia cepacia. As a significant proportion of patients with CF in our community are infected with this organism, we have continued to provide lung transplantation as an option. A retrospective review was conducted of medical records of all patients with CF transplanted between March 1988 and September 1996. Fifty-six transplant procedures were performed in 53 recipients with CF between March 1988 and September 1996. Twenty-eight had B. cepacia isolated pretransplant and 25 remaining positive post-transplant. Of the 53 recipients, 19 have died (15 of 28 [54%] B. cepacia positive and 4 of 25 [16%] B. cepacia negative). B. cepacia was responsible for or involved in 14 deaths. Nine of the deaths occurred in the first 3 mo post-transplantation. One-year survival was 67% for B. cepacia positive patients and 92% for B. cepacia negative patients. Recent modifications in antimicrobial and immunosuppressive therapy since 1995 have resulted in no deaths early post-transplant in the last five patients transplanted. We conclude that early mortality in patients with CF infected with B. cepacia is significantly higher than in those not infected with B. cepacia. Modifications in post-transplant medical therapy may improve outcome.

Background: Depressive symptoms and personality features of patients with mood disorders are supposed to have a strong variability in cross cultural studies. The clinical profile, the outcome and the treatment response seem to be different across the world., Aim: To investigate the differences and similarities of major depressive disorders in Chile and Germany., Material and Methods: Sixty seven Chilean and German depressive inpatients, without comorbidity on axis I or II were studied. Diagnosis of depression was based on DSM IV and ICD 10 criteria for major depression. Symptomatology was assessed using the self reporting questionnaire (SRQ-20), the Symptom checklist-90-revisited (SCL 90-R), the Beck inventory for Depression (BDI), and the Hamilton scale for depression (HAM-D). Personality was assessed with the Munich Personality Scale., Results: Only the SRQ-20 shows significantly higher tendency towards depression among Chilean patients. At the personality level, Chileans exhibited higher scores in extroversion, neuroticism, esoteric and isolating tendencies., Conclusions: These results help to understand differences and similarities between depressive syndromes across cultures. They suggest that the relation between symptomatology, some personality traits and severe major depression has little variability across cultures.

Study Objective: Osteoporosis is a well-recognized complication of lung transplantation that may significantly impair the quality of life of transplant recipients. We performed a prospective study of bone mineral density (BMD) before and after transplantation to determine the degree of bone mass loss associated with lung transplantation Patients and design: We conducted a prospective study of BMD in 28 patients with various end-stage respiratory diseases pretransplantation and 6 to 12 months posttransplantation. The BMD of the lumbar spine (LS) and femoral neck (FN) were measured. All 28 patients were treated only with vitamin D and calcium supplementation posttransplant. The primary endpoint was the percentage change in BMD. The secondary endpoint was the incidence of fractures posttransplant. A univariate analysis was conducted to determine the various risk factors associated with bone mass loss pretransplant and posttransplant., Results: Prior to transplantation, moderate to severe bone disease was evident. The mean (+/- SD) pretransplant T score (the number of SDs from the peak bone mass) and Z score (the number of SDs from the age-matched mean) for the LS were -1.72 +/- 1.37 and -1.44 +/- 1.31, respectively. The mean pretransplant T score and Z score for the FN were -2.65 +/- 1.01 and -1.5 +/- 1.43, respectively. Within 6 to 12 months posttransplant, the mean BMD for the LS decreased by 4.76% (p < 0.001), while the mean BMD for the FN decreased by 5.3% (p < 0.001). Five of the 28 patients (18%) suffered osteoporotic fractures posttransplant, while no fractures were documented pretransplant. The cumulative steroid dose posttransplant was associated with a drop in BMD for the LS and FN (r = 0.39, p = 0.039 and r = 0.63, p < 0.001, respectively), while a negative association was found between cumulative steroid use pretransplant and baseline LS and FN T scores (r = -0.4, p = 0. 02 and r = -0.43, p = 0.023, respectively)., Conclusion: Within 6 to 12 months after lung transplantation, there is a significant decrease in BMD at both the LS and FN levels (approximately 5%) despite vitamin D and calcium supplementation. This drop in BMD is associated with a relatively high incidence of osteoporotic fractures posttransplant.

Background: Immunosuppression and chronic lung disease are known risk factors for mycobacterial infection and might be expected to develop with an increased frequency in lung transplant recipients. We therefore sought to document the incidence and type of mycobacterial infections in a large lung transplant program., Methods: A retrospective review of 219 transplant procedures (60 single lung transplants and 159 double lung transplants) in 210 patients was conducted. All patients had scheduled surveillance bronchoscopies at 3, 6, 9, 12, 18, and 24 months, and yearly thereafter. BAL samples were processed routinely for mycobacterium., Results: Eight patients (3.8%) had evidence of infection (5 men, 3 women; age range, 26 to 63 years). The reasons for transplant were obstructive lung disease (six), cystic fibrosis (one), and pulmonary fibrosis (one). Five recipients had infection in their native lungs; two of five cultured mycobacterium from BAL following transplantation. At least four of five patients had nontuberculous mycobacterium (one showed acid fast bacilli and granuloma on a biopsy specimen that was not sent for culture). None of the five developed disease (mean follow-up = 22 months; range, 3 to 30 months). The organisms were Mycobacterium avium complex (three), Mycobacterium xenopi (one), and unidentified (one). Of the three remaining patients who developed infection after transplantation, one grew Mycobacterium chelonae and the others grew Mycobacterium tuberculosis (both received double lung transplants and had no evidence of mycobacterium in their native lungs). The only definite symptomatic disease occurred in the patients with M tuberculosis, one of whom had evidence of dissemination. The patients with M tuberculosis responded to standard treatment. There have been no deaths due to mycobacterium., Conclusion: Mycobacterial disease rarely occurs following lung transplantation. Cultures for mycobacterium in surveillance BALs in the absence of symptoms are likely unnecessary.

Various factors may influence bioavailability and blood concentrations of cyclosporine, a problem that may be compounded by diseases such as cystic fibrosis in which impaired absorption through the gastrointestinal tract is common. Neoral, a microemulsion formulation of cyclosporine, has improved bioavailability and more stable blood concentrations than earlier formulations. We conducted a prospective, open, crossover study to examine whether these findings held true in 12 clinically stable patients with cystic fibrosis who had undergone lung transplantation at least 6 months earlier. In the first arm, patients continued their usual dosage of cyclosporine twice/day. In the second arm they received Neoral for at least 1 week before having blood studies. For each arm whole blood trough concentrations were drawn for 7-10 successive days, together with a pharmacokinetic study with concentrations drawn at times zero, 1, 2, 3, 4, 6, 12, and 24 hours. Variance was assessed from morning concentrations. Area under the curve from zero to 12 hours (AUC12), maximum concentration (Cmax), and time to Cmax (Tmax) were calculated for each arm. Eleven subjects completed the protocol. The daily variance for Neoral was significantly less than for cyclosporine (p=0.04). The AUC12 for Neoral and cyclosporine were 4164+/-1467 and 5318+/-1670 microg x L/hour (p=0.09), respectively. Respective Cmax were 613+/-242 and 931 +/-458 microg/L (p=0.08) and relative Cmax and AUC12 were 1.91 and 1.47 (p<0.05). Thus Neoral had a superior pharmacokinetic profile and less day-to-day variability in patients with cystic fibrosis who had undergone lung transplantation.

Acute allograft rejection in animals and humans has been associated with increased nitric oxide production in the graft. Exhaled nitric oxide (ENO) measurement is a noninvasive method of assessing inflammation in airway diseases, e.g., asthma, which might be applicable to lung transplant recipients. Over 12 months, ENO of lower respiratory origin was measured in 108 lung transplant recipients with a mean time after transplant of 1,083 d. ENO (mean +/- SEM; ppb) in stable patients (19.5 +/- 1.1; p < 0.001) was not different from that of healthy controls (23.8 +/- 3.2). ENO was significantly higher in episodes of clinical acute rejection (51.1 +/- 6.3) compared with stable patients but not elevated in bronchiolitis obliterans syndrome (18.6 +/- 1.5) or pulmonary infection (25.9 +/- 4.0). A retrospective analysis of bronchoscopy findings and concurrent ENO (n = 99) showed that ENO did not vary according to histological findings (normal, acute rejection grade I, nonspecific inflammatory change) or with a positive BAL culture. ENO was not correlated with differential lymphocyte and neutrophil counts. ENO appears to be a valid marker of clinical acute rejection in human lung transplantation as distinct from infection or bronchiolitis obliterans. Furthermore, bronchoscopic findings in the absence of a clinical illness were not associated with a rise in ENO.

Background: Cytomegalovirus (CMV) disease is an important cause of organ transplant-related morbidity and mortality. During the last 5 years at our institution, prophylactic ganciclovir and hyperimmune globulin have been routinely administered to lung transplant recipients whenever the donor or the recipient was CMV antibody-positive. We sought to assess the efficacy of prophylaxis on viremia, CMV disease, and bronchiolitis obliterans syndrome (BOS)., Methods: A retrospective chart review of 61 consecutive lung transplants performed between recipients between January 1993 and August 1995 was performed. Fifty-six patients who survived at least 1 month were analyzed. Patients were considered at risk for CMV disease whenever pretransplant donor or recipient serology was positive., Results: Fourteen of the 39 patients at risk (36%) had viremia while on prophylaxis. The rate of CMV disease was 13% during the first 6 months following transplantation. A donor whose CMV serology was positive appeared to increase the risk of BOS in a Cox regression model (relative risk=2.4; 95% confidence interval=0.86-6.74; p=0.0957). Neither age, CMV infection (viremia or a positive specimen from BAL), recipient's serology at the time of transplantation, or CMV disease was associated with BOS. None of these variables was associated with mortality on Cox regression analysis or univariate analysis., Conclusions: Administration of combination ganciclovir and hyperimmune globulin prophylactic therapy to lung transplant recipients at risk for CMV infection and disease is associated with a relatively low incidence of disease, which appears only after prophylaxis treatment with ganciclovir is completed. Ganciclovir prophylaxis does not prevent CMV viremia; however, viremia while on prophylaxis is not predictive of disease.

In Maine, prenatal screening for cystic fibrosis (CF) is offered through primary care providers. Cheekbrush (buccal) samples are routinely tested for eight mutations by multiplex PCR amplification of five exons, followed by dot-blot hybridization with pooled allele-specific oligonucleotides (ASO). The ASO methodology is widely used and effective, but somewhat time and labor intensive when applied to CF carrier testing or couple-based prenatal screening in the general pregnant population. Amplification Refractory Mutation System (ARMS) is an improvement of the PCR that allows rapid detection of mutations involving single base changes or small deletions/insertions. In this study, two multiplex ARMS reactions are used to test for 10 common CF mutations. Clinical evaluation of the ARMS test includes a retrospective study of 140 banked samples (54 cell line, proficiency testing, and buccal controls; 86 clinical buccal samples) with known CF genotype (57 with CF mutations, 83 no mutation), followed by a prospective trial in which 309 buccal samples are analyzed con-currently using both methods. The success rate of the ARMS test in buccal lysates is comparable to the ASO method; all CF mutations are successfully identified. For testing nonsterile buccal lysates with low DNA concentrations, optimized performance in the ARMS method is obtained using Amplitaq Gold polymerase. The ARMS method developed is easy, rapid (1 day), and avoids the need for ASO probe labeling, dot-blotting and autoradiography. This study provides further evidence that ARMS methodology is suitable for clinical CF mutation analysis.

Background: Chronic lung allograft rejection manifested by sustained declines in lung function is the most common cause of late death after lung transplantation. Numerous strategies have shown variable results. We sought to evaluate the effect of FK506 (tacrolimus) on bronchiolitis obliterans syndrome (BOS) after lung transplantation., Methods: A single-center open study was conducted of 15 patients whose treatment was converted to tacrolimus from cyclosporine. Of the 15, 12 patients had BOS characterized by sustained loses in lung function while receiving cyclosporine. Rate of decline of forced expiratory volume in 1 second (FEV1) for the 12 patients was calculated before and after administration of tacrolimus. Biochemical changes before and after conversion were compared for the entire group., Results: Median monthly rate of decline in FEV1 was significantly reduced after administration of tacrolimus (5.3% vs 1.1%; p = 0.002). Forced vital capacity did not change significantly. No subjects experienced at least a 10% improvement in FEV1. At least a 10% further decline in FEV1 was noted in five subjects, and seven subjects had no change (i.e., within 10% of baseline). A minor nonsignificant increase in creatinine occurred after administration of tacrolimus. Blood cell count, electrolytes, and liver enzymes remained unchanged. The median change in fasting blood glucose was +0.7 mmol/L (p = 0.02)., Conclusion: Although tacrolimus does not reverse changes in FEV1 with BOS, in this nonrandomized study it seemed to be associated with a decrease in the rate of decline in lung function and no significant sustained toxicity. Further studies are necessary to substantiate this observation.

Advances in surgical technique and better knowledge of the physiologic and immunologic changes in the transplant population, combined with improved diagnostic tools and treatment strategies, have decreased the likelihood of early and, possibly, late mortality caused by a primary infection. Nevertheless, infection continues to be an important cause of death in both the early and late post-transplant periods. Risk of death attributable to infection after prolonged survival, however, is greatest in the setting of chronic rejection. The most significant advances in antimicrobial management have been in the area of prophylaxis. The effectiveness of prophylaxis against P carinii has virtually eliminated that organism as a cause of significant morbidity. Ganciclovir prophylaxis protocols require refinement but have been proved effective against CMV, although that virus continues to be a major pathogen in lung transplant recipients. Ultimately, a careful monitoring protocol and a high index of suspicion for infection requiring investigation and treatment are necessary in the ongoing care of lung transplant recipients. The approach to infections should be guided by the knowledge of the various factors that increase susceptibility to microorganisms and any previous culture and sensitivity results. As transplant physicians try to increase the donor pool through the use of donors who previously might have been rejected and through the potential of xeno-transplantation, vigilance and research must be maintained.

Background: Prolonged survival after lung transplantation is now commonplace as a result of advances in surgical techniques and postoperative management protocols. Although 1- and 5-year functional and survival data after lung transplantation are well known, sparse information is available regarding functional status of recipients surviving beyond 5 years., Methods: The medical records and pulmonary function study results of lung transplant recipients who had survived at least 5 years as of September 1995 were retrospectively reviewed., Results: Of the 76 transplantations performed between November 1983 and September 1990, 30 (39.5%) were double lung transplantations, and 46 (60.5%) were single lung transplantations. Thirty-one recipients were alive 5 years after transplantation (12 double lung transplantations, 19 single lung transplantations). The 5-, 6-, and 7-year survival rates were 44%, 34%, and 29%, respectively. There was no association or difference in cytomegalovirus status, sex, and blood group between those who died within 5 years and those who survived beyond 5 years. The median percent predicted FEVs for single and double lung transplant recipients were as follows: 5 yrs-75%, 75%; 6 years-73%, 75%; 7 years-68%, 73%. The proportion of recipients with bronchiolitis obliterans syndrome according to published criteria was as follows: stage 0, 32%; stage I, 19%; stage II, 16%; and stage III, 19%. The functional status (i.e., active, working, disabled) 5 years after transplantation was as follows: active/working, 74%; active but not working, 13%; some limitation/independent, 10%; and disabled, 3%., Conclusion: Bronchiolitis obliterans syndrome is a frequent occurrence in long-term survivors. Nevertheless, in spite of this condition, most recipients have acceptable lung function, are active, and are generally working.

We reviewed all tissue specimens from 163 transplant patients (108 double lung transplant [DLT], 55 single lung transplant [SLT]) between November 1983 and January 1994 for abnormalities indicating bronchiolitis obliterans organizing pneumonia (BOOP) and found 17 cases (14 DLT and 3 SLT). Of the three SLTs, BOOP was diagnosed by open lung biopsy (OLB) in two and one was found at autopsy. Of the 14 DLTs, BOOP was diagnosed by transbronchial biopsy (TBB) specimens (9), OLB specimens (2), autopsy (1), TBB and OLB specimens (1), and OLB specimens and autopsy (1). BOOP was found between 1 and 43 months posttransplantation; time of survival from diagnosis was between 2 and 36 months with 9 patients presently alive. Concurrent pathologic diagnosis at the time of BOOP findings were as follows: acute rejection (7) (grade 1 [4] and grade 2 [3]), BO and grade 1 rejection (2), BO and grade 2 rejection (2), BO and Aspergillus infection (1), acute alveolar injury (1), acute alveolar injury and pulmonary embolus (1), acute rejection (grade 1) and Burkholderia cepacia pneumonia (1). No other pathologic diagnosis was found in 1 patient. In total, 11 of 17 patients (65%) had associated acute rejection. Of the 17 patients, 7 subsequently developed BO and 3 had BO before the finding of BOOP. Death occurred in 8 patients (5 DLT and the 3 SLT) between 2 and 6 months after the diagnosis. We conclude that BOOP is an important complication after lung transplantation; it was present in 13% of DLTs and 5% of SLTs. BOOP was most often associated with acute rejection.

Pleural complications occurred in 30 (22%) of 138 patients after 53 single and 91 double lung transplants between September 1986 and February 1993. These were defined for the purpose of this study as pneumothorax persisting beyond the first 14 postoperative days, recurrent pneumothorax, or any other pleural process that necessitated diagnostic or therapeutic intervention. Overall, a higher pleural complication rate was seen in double lung transplantation (25 of 30) than in single lung transplantation (5 of 30) with no differences noted in the frequency among preoperative diagnostic groups (p > 0.05). Pneumothorax was the most frequent complication, affecting 14 of 30 patients, with 6 of 14 cases occurring after transbronchial biopsy. All pneumothoraces in single (n = 4) and double lung transplantation (n = 10) resolved spontaneously or with chest tube thoracostomy. One patient required placement of a Clagett window after open lung biopsy and another required thoracotomy and pleural abrasion after transbronchial biopsy. Parapneumonic effusion was observed in 4 of 30 double lung transplantations with spontaneous resolution in all cases. Empyema affected 7 of 30 patients and occurred exclusively in the double lung transplant group. Sepsis developed in three of the patients with this complication and they subsequently died. The risk of empyema was independent of preoperative diagnosis (p > 0.05). Of interest, all patients with cystic fibrosis (n = 3) with complicating empyema had Pseudomonas cepacia in the pleural fluid. Other miscellaneous complications included subpleural hematoma, chylothorax, and hemothorax. The latter two necessitated thoracic duct and bronchial artery ligation, respectively. In summary, a significant proportion of lung transplant recipients will have pleural space complications. The vast majority of these will resolve spontaneously or with conservative procedures. These complications were not related to preoperative diagnosis nor associated with a significant prolongation of hospital stay (p > 0.05). Empyema is the only pleural space complication associated with increased patient mortality and, as such, is an important clinical marker for those at risk for sepsis and death.

After early experience with perioperative bleeding in sequential single-lung transplant recipients, aprotinin was introduced in an attempt to reduce this complication and the attendant morbidity. Records of sequential single-lung transplantations (n = 33) performed between January 1989 and November 1991 were reviewed to assess the impact of aprotinin on perioperative blood loss and blood product requirements. Recipients were divided according to whether or not they required cardiopulmonary bypass. In patients requiring cardiopulmonary bypass (n = 15), mean estimated postoperative blood loss was 3,000 +/- 500 mL in those who did not receive aprotinin (n = 4) compared with 1,177 +/- 253 mL in those who received aprotinin (n = 11) (p < 0.05). An average of 8.0 +/- 0.7 units of packed red blood cells were administered to patients not receiving aprotinin compared with 3.1 +/- 0.7 units to those who received aprotinin (p < 0.05). Requirements for fresh frozen plasma were similar in each group. There were no differences in blood loss or blood product replacement in the group not undergoing cardiopulmonary bypass (n = 18). Therefore, we conclude that aprotinin decreases postoperative blood loss and blood product requirements in patients undergoing sequential single-lung transplantation under cardiopulmonary bypass.

Between November 1983 and August 1993, The Toronto Lung Transplant Program performed 153 transplantations in 144 recipients: 53 single-lung transplantations (SLT) and 100 double-lung transplantations (DLT). Thirty-eight open lung biopsies (OLBs) were done in 32 (22% of all recipients): 19 in SLT (36% of SLT) 12 in DLT (12% of DLT), and 1 in a patient who had a SLT and then a double retransplantation. Six recipients underwent OLB twice: 1 DLT, 3 SLT, and 2 who had OLB both before and after retransplantation. Indication for 11 early OLBs (< or = 45 days postoperative) was persistent parenchymal infiltrates. Indications for 27 late OLBs (> 45 days postoperative) included progressive radiologic disease with clinical findings or progressive loss of pulmonary function (18), persistent poor graft function (3), mass or nodules (3), persistent infiltrates without functional loss (2), and persistent lymphocytosis in bronchoalveolar lavage (1). Open lung biopsy confirmed a previous clinical or pathologic diagnosis in 11, suggested a diagnosis in 2, yielded nonspecific information in 16, and provided different diagnosis in 9. New diagnosis that changed therapy was made in 1 of 11 early OLBs and in 8 of 27 late OLBs. These 9 diagnoses included in SLTs: bronchiolitis obliterans (2), bronchiolitis obliterans organizing pneumonia (1), malignant lymphoma (1), and chronic vascular rejection (1) in SLT, and bronchiolitis obliterans organizing pneumonia (3) and Burkholderia cepacia infection (1) in DLT. We conclude that OLB is of little value in the perioperative period but yields useful information in approximately 30% of patients when performed late.

Background: Acute alveolar injury is a pathologic description of a nonspecific response of the lung to an acute injury. It has been our experience that acute alveolar injury may be seen in transbronchial biopsy and open lung biopsy specimens from patients who have successfully undergone lung transplantation. Because no studies of the clinical significance of acute alveolar injury in lung transplant recipients are available in the literature, we reviewed the impact of this pathologic finding in patients who underwent transplantation in the Toronto Lung Transplant Program., Methods: From November 1983 through December 1992, we retrospectively reviewed all transbronchial biopsies and open lung biopsies performed in lung transplant recipients., Results: Of the 137 transplantations performed (53 single and 84 double) acute alveolar injury was observed in 21 single (34 of 173 transbronchial biopsy and 3 of 11 open lung biopsy) lung biopsy) and 22 double (38 of 415 transbronchial biopsy and 3 of 11 open lung biopsy) lung transplantations. We sought to explain this finding on the basis of the concurrent clinical scenario. Acute alveolar injury occurred most commonly in association with infection (52%) followed by postoperative period (19%) and acute (16%) rejection. Acute alveolar injury occurred as an isolated finding in 7% of patients. It occurred within the first 4 months after transplantation in 80% of cases. In 10 of 21 single lung transplantations and 12 of 22 double lung transplantations, acute alveolar injury was seen at least twice. However, no difference was found in mortality between such patients (6 of 10 single and 6 of 12 double lung transplantations) and patients with only one episode of acute alveolar injury., Conclusion: Acute alveolar injury is a relatively common finding in histologic specimens from patients with lung transplantation and is most commonly associated with infection.

We investigated the utility of transbronchial lung biopsy in allograft evaluation, particularly with reference to rejection-mediated bronchiolitis obliterans in 105 transplant recipients surviving 3 months or more (29 single lung, 76 double lung). A sensitivity and specificity of one transbronchial biopsy procedure with an average procurement of 7.6 tissue fragments was 17.1% and 94.5%, respectively, according to results obtained from biopsies carried out after 2 months transplantation in 29 patients with confirmed rejection-mediated bronchiolitis obliterans and 61 patients known not to have the disease. The predictive value of a positive procedure for the presence of disease was 65.5% and that of a negative procedure for the absence of disease was 65.2%. Similarly, the sensitivity and specificity for the finding of bronchiolar mural fibrosis were 18.5% and 85.3%, respectively; and, for lymphocytic bronchiolitis, the values were 1.9% and 100%, respectively. We conclude that, although the sensitivity of transbronchial biopsy and the predictive value of a procedure which shows microscopic bronchiolitis obliterans are low, attempts should be made to support a clinical diagnosis of bronchiolitis obliterans through biopsy, given the problems inherent in achieving an early and certain diagnosis according to clinical criteria alone.

Between November 1983 and September 1992, The Toronto Lung Transplant Program performed 131 lung transplantations in 122 recipients; 53 single lung transplantations and 78 double lung transplantations. Forty-five patients died, 25 (47%) in the single lung transplantation and 20 (25%) in the double lung transplantation groups. We retrospectively reviewed the hospital charts of all deceased recipients and the postmortem reports of the 35 patients (20 single lung transplantations and 15 double lung transplantations) who had autopsies. Preoperative single lung transplantation diagnoses included pulmonary fibrosis, (n = 17) obstructive disease (n = 6) and vascular disease (n = 2). Preoperative diagnosis of double lung transplantation included pulmonary fibrosis (n = 2), obstructive disease (n = 6), septic lung disease (n = 9), and vascular disease (n = 3). The most common cause of death in single lung transplantation was infection. Five patients died of bronchiolitis obliterans, and five more had bronchiolitis obliterans lesions present at autopsy that were not a direct cause of death. Diagnosis of primary disease was made in 23 of 25 single lung transplantations antemortem and 2 of 25 at autopsy. Autopsy diagnoses were disseminated Aspergillus and cytomegalovirus infection. In double lung transplantations, infection was also the primary cause of death; in three other patients, airway dehiscence preceded infection. Bronchiolitis obliterans was the second most common cause of death and was also present in four patients dying of infection. All double lung transplantation diagnoses were made antemortem. We concluded that infection and then bronchiolitis obliterans are the primary causes of death after lung transplantation. Although infection is a major cause both early and late after transplantation, bronchiolitis obliterans is an important factor in transplantation only late after the operation.

We previously identified a polymorphism in the human estrogen receptor (ER) gene, within the coding region for the protein's amino terminal B-domain. In estrogen receptor-positive (ER+) breast tumors, the variant allele was preferentially associated with lower levels of ER, and was clinically correlated with frequent spontaneous abortions. DNA sequencing revealed a point mutation that changes codon 86 from Ala to Val and a silent mutation in codon 87. Because we initially detected the variant allele by analyzing RNA, only those tissues in which the ER gene is actively expressed were suitable for genotype analysis. We now describe an assay that uses genomic DNA as the substrate for determining the ER B genotype, DNA containing the polymorphic region of the ER gene is amplified by the polymerase chain reaction, then the amplified DNA is hybridized with radiolabeled oligonucleotide probes complementary to the wild type and variant ER alleles. This method allowed us to determine the ER B genotype of women with ER+ and ER- tumors, starting with minute amounts of DNA from frozen or paraffin embedded tissues. ER B genotyping was also performed on women without breast cancer using DNA extracted from blood cells. The combined results from analyses of RNA and DNA from 300 breast cancer patients showed that 12% were heterozygotes. In the ER+ group (n = 183), 11.5% carried the variant gene compared to 12.8% in the ER-negative group (n = 117) (chi 2 = 0.11; df = 1; p greater than 0.25).(ABSTRACT TRUNCATED AT 250 WORDS)