An evaluation of methods for the isolation of nontuberculous mycobacteria from patients with cystic fibrosis, bronchiectasis and patients assessed for lung transplantation.

BACKGROUND: RGM medium is an agar-based, selective culture medium designed for the isolation of nontuberculous mycobacteria (NTM) from the sputum of patients with cystic fibrosis (CF). We evaluated RGM medium for the detection of NTM in patients with CF (405 samples), bronchiectasis (323 samples) and other lung diseases necessitating lung transplantation (274 samples). METHODS: In total, 1002 respiratory samples from 676 patients were included in the study. Direct culture on RGM medium, with incubation at two temperatures (30 °C and 37 °C), was compared with conventional culture of decontaminated samples for acid-fast bacilli (AFB) using both a solid medium (Löwenstein-Jensen medium) and a liquid medium (the Mycobacterial Growth Indicator Tube; MGIT). RESULTS: For all three patient groups, significantly more isolates of NTM were recovered using RGM medium incubated at 30 °C than by any other method (sensitivity: 94.6% vs. 22.4% for conventional AFB culture; P < 0.0001). Significantly more isolates of Mycobacterium abscessus complex were isolated on RGM at 30 °C than by AFB culture (sensitivity: 96.1% vs. 58.8%; P < 0.0001). The recovery of Mycobacterium avium complex was also greater using RGM medium at 30 °C compared to AFB culture (sensitivity: 83% vs. 70.2%), although this difference was not statistically significant and a combination of methods was necessary for optimal recovery (P = 0.21). CONCLUSIONS: In the largest study of RGM medium to date, we reaffirm its utility for isolation of NTM from patients with CF. Furthermore; we show that it also provides an effective tool for culture of respiratory samples from patients with bronchiectasis and other lung diseases.

Maintaining surgical skills for military general surgery: the potential role for multivisceral organ retrieval in military general surgery training and practice.

The closure of the Medical Treatment facility in Camp BASTION and the return to contingency operations presents a new challenge in training and maintaining the skills of military surgeons. Multivisceral organ retrieval presents a unique opportunity to practice some of the more unusual techniques required in military surgery in the National Health Service. This article details the experience that organ retrieval offers and matches this to the needs of military surgeons. National Organ Retrieval Service teams need skilled surgeons, and a mutually beneficial partnership is in prospect.

Raised interleukin-17 is immunolocalised to neutrophils in cystic fibrosis lung disease.

Interleukin (IL)-17 is pivotal in orchestrating the activity of neutrophils. Neutrophilic inflammation is the dominant pathology in cystic fibrosis (CF) lung disease. We investigated IL-17 protein expression in the lower airway in CF, its cellular immunolocalisation and the effects of IL-17 on CF primary bronchial epithelial cells. Immunohistochemistry was performed on explanted CF lungs and compared with the non-suppurative condition pulmonary hypertension (PH). Airway lavages and epithelial cultures were generated from explanted CF lungs. Immunoreactivity for IL-17 was significantly increased in the lower airway epithelium in CF (median 14.1%) compared with PH (2.95%, p=0.0001). The number of cells staining positive for IL-17 in the lower airway mucosa was also increased (64 cells·mm(-1) compared with 9 cells·mm(-1) basement membrane, p=0.0005) and included both neutrophils in addition to mononuclear cells. IL-17 was detectable in airway lavages from explanted CF lungs. Treatment of epithelial cultures with IL-17 increased production of IL-8, IL-6 and granulocyte macrophage colony-stimulating factor. In conclusion, immunoreactive IL-17 is raised in the lower airway of people with CF and localises to both neutrophils and mononuclear cells. IL-17 increases production of pro-neutrophilic mediators by CF epithelial cells, suggesting potential for a positive feedback element in airway inflammation.

Liver resection for colorectal cancer metastases involving the caudate lobe.

BACKGROUND: Up to 5 per cent of liver resections for colorectal cancer metastases involve the caudate lobe, with cancer-involved resection margins of over 50 per cent being reported following caudate lobe resection. METHODS: Outcomes of consecutive liver resections for colorectal metastases involving the caudate lobe between 1996 and 2009 were reviewed retrospectively, and compared with those after liver surgery without caudate resection. RESULTS: Twenty-five patients underwent caudate and 432 non-caudate liver resection. Caudate resection was commonly performed as part of extended resection. There were no differences in operative complications (24 versus 21·1 per cent; P = 0·727) or blood loss (median 300 versus 250 ml; P = 0·234). The operating time was longer for caudate resection (median 283 versus 227 min; P = 0·024). Tumour size was larger in the caudate group (median 40 versus 27 mm; P = 0·018). Resection margins were smaller when the caudate lobe was involved by tumour, than in resections including tumour-free caudate or non-caudate resection; however, there was no difference in the proportion of completely excised tumours between caudate and non-caudate resections (96 versus 96·1 per cent; P = 0·990). One-year overall survival rates were 90 and 89·3 per cent respectively (P = 0·960), with 1-year recurrence-free survival rates of 62 and 71·2 per cent (P = 0·340). CONCLUSION: Caudate lobe surgery for colorectal cancer liver metastases does not increase the incidence of resection margin involvement, although when the caudate lobe contains metastases the margins are significantly closer than in other resections.

Inflammation and epithelial to mesenchymal transition in lung transplant recipients: role in dysregulated epithelial wound repair.

Epithelial to mesenchymal transition (EMT) has been implicated in the pathogenesis of obliterative bronchiolitis (OB) after lung transplant. Although TNF-alpha accentuates TGF-beta1 driven EMT in primary human bronchial epithelial cells (PBECs), we hypothesized that other acute pro-inflammatory cytokines elevated in the airways of patients with OB may also accentuate EMT and contribute to dysregulated epithelial wound repair. PBECs from lung transplant recipients were stimulated with TGF-beta1+/-IL-1beta, IL-8, TNF-alpha or activated macrophages in co-culture and EMT assessed. The quality and rate of wound closure in a standardized model of lung epithelial injury was assessed in response to above stimuli. Co-treatment with TGF-beta1+TNF-alpha or IL-1beta significantly accentuates phenotypic and some functional features of EMT compared to TGF-beta1 alone. Co-treatment with TGF-beta1+TNF-alpha or IL-1beta accelerates epithelial wound closure however the quality of repair is highly dysregulated. Co-treatment with TGF-beta1+IL-8 has no significant effect on EMT or the speed or quality of wound healing. Activated macrophages dramatically accentuate TGF-beta1-driven EMT and cause dysregulated wound repair. Crosstalk between macrophage-derived acute inflammation in the airway and elevated TGF-beta1 may favor dysregulated airway epithelial repair and fibrosis in the lung allograft via EMT.

Bronchial epithelial cells cultured from clinically stable lung allograft patients promote the development of macrophages from monocytes rather than dendritic cells.

BACKGROUND: It is understood that chronic allograft failure occurs as a result of alloimmune and non-alloimmune injury. Dendritic cells (DC) are thought to be crucial in regulating (allo)immune airway damage and interactions with epithelial cells are likely. Studies in human lung transplantation are limited, however, and the available literature on DC is inconsistent. This study focused on the ex vivo influence of primary bronchial epithelial cells derived from lung allografts on DC differentiation. METHODS: Epithelial cell conditioned media (ECCM) were added to monocytes differentiating into DC under the influence of interleukin-4 and granulocyte macrophage-colony stimulating factor. The resultant cells were compared with DC cultured without ECCM and with monocyte-derived macrophages. Expression of typical DC (eg, CD1a) and macrophage (eg, CD14) markers was assessed by flow cytometry. Phenotypical assessments were complemented by functional studies of mannose receptor-mediated phagocytosis (FITC-dextran uptake) and antigen-presenting capability (mixed lymphocyte reactions). RESULTS: Cells exposed to ECCM expressed significantly lower levels of CD1a than unexposed DC. CD14 expression and phagocytic function were increased. ECCM cultured cells also expressed lower levels of T cell co-stimulatory molecules, secreted an anti-inflammatory cytokine profile and had significantly reduced antigen-presenting capability. CONCLUSION: Using phenotypic and functional approaches, this study has shown that ECCM from lung allografts drives the production of macrophage-like cells from monocytes rather than DC. The data suggest that epithelial cells may restrain airway DC and potential alloimmunity. It is unclear whether the observed effect is specifically seen in lung transplant recipients or is a general property of bronchial epithelial cells. This may reflect a homeostatic inter-relationship between airway epithelial and DC populations relevant both to lung allografts and the lung more generally.

Epithelial to mesenchymal transition (EMT) and airway remodelling after human lung transplantation.

BACKGROUND: Aberrant epithelial repair is a key event in the airway remodelling which characterises obliterative bronchiolitis (OB) in the transplanted lung. The potential for airway epithelium from lung transplant recipients to undergo epithelial to mesenchymal cell transition (EMT) was assessed in culture and in vivo in lung allograft tissue. METHODS: Change in epithelial and mesenchymal marker expression was assessed after stimulation with transforming growth factor beta(1) (TGF-beta(1)) alone or in combination with tumour necrosis factor alpha (TNFalpha) and compared with untreated controls. The ability of cells to deposit extracellular matrix, secrete matrix metalloproteinases (MMPs) and invade collagen was investigated. Immunolocalisation of epithelial and mesenchymal markers was compared in airway tissue from stable recipients and those with OB. RESULTS: Untreated cells maintained epithelial morphology and phenotype. TGF-beta(1) reduced expression of epithelial markers, increased expression of vimentin and fibronectin, promoted collagen I and fibronectin deposition and increased MMP-9 production. Co-treatment with TNFalpha dramatically accentuated phenotypic and some functional features of EMT. Airway epithelial biopsies from recipients with OB demonstrated significantly increased staining for mesenchymal markers and significantly reduced E-cadherin staining compared with stable recipients. CONCLUSIONS: These observations demonstrate the ability of human airway epithelium to undergo EMT and suggest this phenomenon may be a potential link between inflammatory injury and TGF-beta(1)-driven airway remodelling in the development of OB.

Airway epithelial cell senescence in the lung allograft.

Chronic lung allograft dysfunction, manifesting as bronchiolitis obliterans syndrome (BOS), is characterized by airway epithelial injury, impaired epithelial regeneration and subsequent airway remodeling. Increased cellular senescence has been reported in renal and liver allografts affected by chronic allograft dysfunction but the significance of cellular senescence in the airway epithelium of the transplanted lung is unknown. Thirty-four lung transplant recipients, 20 with stable graft function and 14 with BOS, underwent transbronchial lung biopsy and histochemical studies for senescence markers in small airways. Compared to nontransplant control lung tissue (n = 9), lung allografts demonstrate significantly increased airway epithelial staining for senescence-associated beta galactosidase (SA beta-gal) (p = 0.0215), p16(ink4a) (p = 0.0002) and p21(waf1/cip) (p = 0.0138) but there was no difference in expression of these markers between stable and BOS affected recipients (p > 0.05). This preliminary cross-sectional study demonstrates that cellular senescence occurs with increased frequency in the airway epithelium of the lung allograft but does not establish any association between airway epithelial senescence and BOS. A prospective longitudinal study is required to better address any potential causal association between airway epithelial senescence in stable allograft recipients and the subsequent development of BOS.

Outcomes of lung transplantation for cystic fibrosis in a large UK cohort.

BACKGROUND: Lung transplantation is an important option to treat patients with advanced cystic fibrosis (CF) lung disease. The outcomes of a large UK cohort of CF lung transplantation recipients is reported. METHODS: Retrospective review of case notes and transplantation databases. RESULTS: 176 patients with CF underwent lung transplantation at our centre. The majority (168) had bilateral sequential lung transplantation. Median age at transplantation was 26 years. Diabetes was common pretransplantation (40%). Polymicrobial infection was common in individual recipients. A diverse range of pathogens were encountered, including the Burkholderia cepacia complex (BCC). The bronchial anastomotic complication rate was 2%. Pulmonary function (forced expiratory volume in 1 s % predicted) improved from a pretransplantation median of 0.8 l (21% predicted) to 2.95 l (78% predicted) at 1 year following transplantation. We noted an acute rejection rate of 41% within the first month. Our survival values were 82% survival at 1 year, 70% at 3 years, 62% at 5 years and 51% at 10 years. Patients with BCC infection had poorer outcomes and represented the majority of those who had a septic death. Data are presented on those free from these infections. Bronchiolitis obliterans syndrome (BOS) and sepsis were common causes of death. Freedom from BOS was 74% at 5 years and 38% at 10 years. Biochemical evidence of renal dysfunction was common although renal replacement was infrequently required (<5%). CONCLUSION: Lung transplantation is an important therapeutic option in patients with CF even in those with more complex microbiology. Good functional outcomes are noted although transplantation associated morbidities accrue with time.

Antimicrobial peptides in lung transplant recipients with bronchiolitis obliterans syndrome.

Mechanisms other than classical alloimmunity are implicated in the pathogenesis of bronchiolitis obliterans syndrome (BOS). It was hypothesised that antimicrobial peptides (AMPs), elements of the innate immune response, have a role in BOS pathogenesis. Pulmonary expression of the neutrophil-derived AMPs human cathelicidin (hCAP)-18/LL-37 and alpha-defensins (human neutrophil peptides (HNP) 1-3), and the epithelial cell-derived AMPs human beta-defensin (hBD)-2, elafin and secretory leukoprotease inhibitor (SLPI) were measured in stable lung transplant recipients and those with BOS. The relationship between airway pathogens and AMP levels was examined. Bronchoalveolar lavage (BAL) was performed on 44 lung transplant recipients (30 stable, 14 with BOS). BAL was cultured for pathogens and ELISA for AMPs was performed. The presence of airway pathogens was associated with significantly increased levels of neutrophil-derived and epithelial-derived AMPs. When patients without pathogens in BAL fluid were analysed, eight recipients with BOS had elevated hCAP-18/LL-37 and HNP 1-3 compared with 25 stable recipients. hBD-2 and elafin levels were comparable in BOS and stable recipients, but SLPI levels were reduced in BOS. Bronchiolitis obliterans syndrome is associated with elevated airway human cathelicidin 18/LL-37 and human neutrophil peptides 1-3 from activated neutrophils, even in the absence of pathogens. Together with reduced airway secretory leukoprotease inhibitor this may favour nonalloimmune airway injury with reduced antiprotease defence and increased neutrophil degranulation.

Does the laparoscopic colorectal surgery learning curve adversely affect the results of colorectal cancer resection? A 3-year prospective study in a district general hospital.

INTRODUCTION: Laparoscopic colorectal surgery is slowly being adopted across the UK. We present a 3-year prospective study of laparoscopic colorectal cancer resections in a district general hospital. METHOD: Data relating to premorbid, operative and postoperative parameters were recorded for all patients undergoing laparoscopic, open, planned converted (laparoscopic assisted) and unplanned converted resections prospectively from April 2003 to April 2006. RESULTS: A total of 238 colorectal resections were performed, 153 of which were for cancer. Of these 44 (29%) were open, 77 (50%) were laparoscopic and 32 (21%) were converted [26 (17%) planned and six (4%) unplanned]. Blood loss was less in the laparoscopic group compared with the open group (P = 0.02) as was intra-operative fluid replacement (P = 0.01). Time to requiring oral analgesia alone was shorter (P = 0.001) and bowel function returned earlier (P = 0.001) in the laparoscopic group. This is reflected in a trend towards a shorter hospital stay for the laparoscopic group compared with the open group (P = 0.049). The operating time of the laparoscopic group was not significantly longer (P = 0.38). The complication rate was similar between groups (P = 0.31) and the mortality in the laparoscopic group was 1.3%. CONCLUSION: Changing from open to laparoscopic dissection for colorectal cancer is safe even during the initial learning curve. There are clear potential short-term benefits for patients and the technique can be introduced without penalties in terms of reduced surgical throughput.

Morgagni hernia presenting with bowel obstruction in a lung transplant recipient: case report.

Morgagni hernias are uncommon congenital diaphragmatic deficiencies that may remain asymptomatic till adulthood. We report a case of Morgagni hernia presenting with subacute bowel obstruction in a bilateral lung transplant recipient. This diaphragmatic deficiency was not evident during bilateral lung transplantation surgery via clamshell incision. To our knowledge this is the first report of a congenital defect evident after lung transplantation.

A qualitative synthesis of gastro-oesophageal reflux in bronchiectasis: Current understanding and future risk.

Gastro-oesophageal reflux disease (GORD) is a common comorbidity in bronchiectasis, and is often associated with poorer outcomes. The cause and effect relationship between GORD and bronchiectasis has not yet been fully elucidated and a greater understanding of the pathophysiology of the interaction and potential therapies is required. This review explores the underlying pathophysiology of GORD, its clinical presentation, risk factors, commonly applied diagnostic tools, and a detailed synthesis of original articles evaluating the prevalence of GORD, its influence on disease severity and current management strategies within the context of bronchiectasis. The prevalence of GORD in bronchiectasis ranges from 26% to 75%. Patients with co-existing bronchiectasis and GORD were found to have an increased mortality and increased bronchiectasis severity, manifest by increased symptoms, exacerbations, hospitalisations, radiological extent and chronic infection, with reduced pulmonary function and quality of life. The pathogenic role of Helicobacter pylori infection in bronchiectasis, perhaps via aspiration of gastric contents, also warrants further investigation. Our index of suspicion for GORD should remain high across the spectrum of disease severity in bronchiectasis. Identifying GORD in bronchiectasis patients may have important therapeutic and prognostic implications, although clinical trial evidence that treatment targeted at GORD can improve outcomes in bronchiectasis is currently lacking.

Observational Study of Methotrexate in the Treatment of Bronchiolitis Obliterans Syndrome.

BACKGROUND: Methotrexate (MTX) is potential change in immunosuppression after lung transplantation that may help to slow down the decline in lung function in bronchiolitis obliterans syndrome (BOS). METHODS: We sought to analyze the safety and efficacy of MTX in patients with BOS, by retrospective case review. RESULTS: Thirty lung allograft patients were treated with MTX for BOS after one bilateral lower lobe, nine single, 16 bilateral, and four heart-lung transplants. Twenty-one patients had MTX treatment for a minimum of 6 months, and their serial lung function was analyzed for efficacy. In these patients, there was a significant overall increase in mean forced expiratory volume in 1 second (FEV1) of 149 mL (P < .02) at 3 months, with an increase observed in 14 of 21 patients. At 6 months, there was a mean increase in FEV1 of 117 mL (P < .05). At 12 months, there was a mean non-significant increase of FEV1 of 60 mL (P = .19) observed in 18 patients who had MTX for this time period. The rate of decline in FEV1 before MTX was 118.5 mL/month and at 3 months after MTX increased to 49.5 mL/months (P < .0005) in the FEV1. Nine patients had been treated with MTX for less than 6 months; two died within 6 months of starting MTX, five tolerated the drug poorly with nausea and tiredness, and one developed leucopenia. One patient requested discontinuation of the medication after failing to halt the rapid progressive decline in lung function after 1 month. CONCLUSIONS: Methotrexate therapy provides a potential therapeutic strategy in managing the progressive decline in lung function observed in BOS. This is hampered by the observation of poor tolerability and side effects.

Hypothesis: epithelial-to-mesenchymal transition is a common cause of chronic allograft failure.

Renal, hepatic, and lung allografts are compromised by aggressively deteriorating function. This chronic process is produced by an overall burden of organ damage, but the pathophysiology remains poorly understood. Rates of chronic rejection in the lung, for example, have not substantially improved over the last decade, despite new immunosuppressive drugs and improvements in surgical procedure. We present a hypothesis that epithelial-to-mesenchymal transition is a common cause of chronic allograft failure. Research in this area may provide insights into chronic rejection of kidney, liver, and lung allografts that impact on future therapeutic strategies.

The Interaction between Diabetes, Body Mass Index, Hepatic Steatosis, and Risk of Liver Resection: Insulin Dependent Diabetes Is the Greatest Risk for Major Complications.

Background. This study aimed to assess the relationship between diabetes, obesity, and hepatic steatosis in patients undergoing liver resection and to determine if these factors are independent predictors of major complications. Materials and Methods. Analysis of a prospectively maintained database of patients undergoing liver resection between 2005 and 2012 was undertaken. Background liver was assessed for steatosis and classified as <33% and ≥33%. Major complications were defined as Grade III-V complications using the Dindo-Clavien classification. Results. 504 patients underwent liver resection, of whom 56 had diabetes and 61 had steatosis ≥33%. Median BMI was 26 kg/m(2) (16-54 kg/m(2)). 94 patients developed a major complication (18.7%). BMI ≥ 25 kg/m(2) (P = 0.001) and diabetes (P = 0.018) were associated with steatosis ≥33%. Only insulin dependent diabetes was a risk factor for major complications (P = 0.028). Age, male gender, hypoalbuminaemia, synchronous bowel procedures, extent of resection, and blood transfusion were also independent risk factors. Conclusions. Liver surgery in the presence of steatosis, elevated BMI, and non-insulin dependent diabetes is not associated with major complications. Although diabetes requiring insulin therapy was a significant risk factor, the major risk factors relate to technical aspects of surgery, particularly synchronous bowel procedures.