André Dutly

OCT4 expression in human non-small cell lung cancer: implications for therapeutic intervention

Here we investigate the expression of OCT4 human lung adenocarcinoma and bronchioloalveolar carcinoma (BAC) tumor biopsies and tumor-derived primary cell cultures. OCT4 has been detected in several human tumors suggesting a potentially critical role in tumorigenesis. We assessed the presence of OCT4 in clinical tumor samples of both adenocarcinoma and BAC at the cellular and transcriptional levels, respectively. Furthermore, we evaluated tumor-derived cell cultures for potential differences in OCT4 expression. Immunohistochemical analysis depicted OCT4 in 2 of 8 adenocarcinoma tumor samples and 3 of 5 BAC tumor samples, with no apparent difference in the degree of expression among the sections examined. These results were validated by transcript analysis. Flow cytometric assessment of 11 adenocarcinoma-derived cell cultures and 3 BAC-derived cell cultures revealed significantly higher OCT4 expression in adenocarcinoma tumors compared to their normal counterparts. This, however, was not observed in the BAC cultures. Comparative studies of OCT4 in adenocarcinoma and BAC tumor cell cultures demonstrated a dramatically higher expression in the former. The expression of OCT4 may represent a specific and effective target for therapeutic intervention in adenocarcinoma and BAC. In addition, the aberrant expression and distribution of OCT4 may indicate important parameters concerning the differences between adenocarcinoma and BAC.

Melatonin attenuates posttransplant lung ischemia-reperfusion injury

BACKGROUND Melatonin, a pineal hormone, is a free radical scavenger and an antioxidant. The purpose of this study was to assess the protective effect of melatonin on posttransplant lung ischemia-reperfusion injury. METHODS Rat single-lung transplantation was performed in two (n = 10) experimental groups after 18 hours of cold (4 degrees C) ischemia. Group I animals consisted of the ischemic control group. In group II, donor and recipient animals were treated with intraperitoneal injection of 10 mg/kg melatonin 10 minutes before harvest and reperfusion, respectively. After 2 hours of reperfusion, oxygenation, plasma, and bronchoalveolar lavage nitrite levels were measured. Lung tissue was assessed for thiobarbituric acid reactive substances and myeloperoxidase activity. Peak airway pressure was recorded throughout the reperfusion period. RESULTS The melatonin-treated group showed significantly better oxygenation (321.8+/-33.8 mm Hg versus 86.1+/-17.4 mm Hg; p < 0.001), reduced lipid peroxidation (0.65+/-0.3 nmol/g versus 1.63+/-0.8 nmol/g; p = 0.032), and reduced myeloperoxidase activity (0.56+/-0.1 deltaOD x mg(-1) x min(-1) versus 1.01+/-0.2 deltaOD x mg(-1) x min(-1); p = 0.032). Bronchoalveolar lavage nitrite levels in the transplanted lungs were significantly lower in group II than in group I (0.34+/-0.06 micromol/L versus 1.65+/-0.6 micromol/L; p = 0.016). In group II significant reduction in peak airway pressure was noted compared with group I (p = 0.002). CONCLUSIONS In this model, exogenously administered melatonin effectively protected lungs from reperfusion injury after prolonged ischemia.

Accelerated treatment for early and late postpneumonectomy empyema

BACKGROUND Postpneumonectomy empyema is a rare but serious complication of pneumonectomy. Despite use of various therapeutic approaches and techniques during the last five decades, successful therapy remains difficult and is often associated with high morbidity and prolonged hospitalization. METHODS We evaluated a concept for accelerated treatment, which consists of radical debridement of the pleural cavity and packing with wet dressings of povidoneiodine. This was repeated in the operating theater every second day, until the chest cavity was macroscopically clean. If present, bronchial stump insufficiency was closed and secured by omentopexy. Finally, the pleural space was obliterated with antibiotic solution. RESULTS Twenty patients, 13 with early postpneumonectomy empyema (10 to 89 days; mean, 37 days) and 7 with late postpneumonectomy empyema (124 to 7,200 days; mean, 1,126 days) were treated. Fifteen patients presented with bronchopleural fistula (11 right, 4 left), which developed after chemotherapy (n = 6) or after radiotherapy (n = 3) (unknown cause in 4 patients). Six patients were referred after previously unsuccessful surgical attempts. Pleural cultures were positive in 17 cases for one or several bacteria including fungoides (n = 2). The average number of interventions was 3.5 (3 to 5). The chest was definitively closed in all patients within 8 days. Mean hospitalization time was 17 days (7 to 35 days). During the same hospitalization, 2 patients needed reoperation because of an undetected bronchopleural fistula. Postpneumonectomy empyema was successfully treated in all patients. There was no in-hospital or 3-month postoperative mortality. CONCLUSIONS Repeated surgical debridement combined with closure of bronchopleural fistula and antimicrobial therapy enables successful treatment of early and late postpneumonectomy empyema within a short period and is a well-tolerated concept.

Thoracoscopic lobectomy for benign disease – a single centre study on 64 cases

OBJECTIVE Chronic lung infection is the main indication for lobectomy in benign pulmonary disease and may be technically demanding due to inflammatory changes such as adhesions, lymph node enlargement and neovascularization. The role of the thoracoscopic operation in these indications is yet ill-defined. METHODS We retrospectively analyzed the results of patients who underwent thoracoscopic lobectomy (TL) between 1992 and June 1999 and compared this study group with patients who underwent open lobectomy (OL), all for benign disease. Data were not normally distributed, therefore, the median and range is given and nonparametric statistical analysis was applied. RESULTS A total of 117 lobectomies for benign disease (64 TL) were analyzed. Indications included bronchiectasis (36 TL; 18 OL), chronic infections (13 TL; eight OL), tuberculosis (five TL; 15 OL), emphysema (five TL; one OL), AV-malformations (two TL; one OL), severe haemoptysis (four OL), and others (three TL; six OL). Twelve conversions to thoracotomy were necessary due to severe adhesions. One patient in the open lobectomy group died within 30 days postoperative. Drainage time was 5.0 (1-32) days in TL and 6.0 (3-21) days in OL, hospital stay was 8.5 (4-41) days and 10.0 (5-52) days, respectively. Blood loss was 0 (0-2000) ml in TL and 300 (0-6000) ml in OL. Operation time for thoracoscopic lobectomies significantly decreased from 2.5 (1-6) h for cases between 1992 and 1997 (n=49) to 1.5 (0.5-2.5) h for recent cases (n=15) (P<0.01). In addition, a trend towards less blood loss was noted (100 (0-2000) ml vs. 0 (0-400) ml; P=0.06). Drainage time and hospital stay did not differ significantly. CONCLUSIONS Thoracoscopic lobectomy in chronic inflammatory disease can be performed safely in selected patients, especially with bronchiectasis. Conversion rate to thoracotomy is low. Operation time with this approach declined significantly over time.

Identification of mesenchymal stromal cells in human lung parenchyma capable of differentiating into aquaporin 5-expressing cells

The lack of effective therapies for end-stage lung disease validates the need for stem cell-based therapeutic approaches as alternative treatment options. In contrast with exogenous stem cell sources, the use of resident progenitor cells is advantageous considering the fact that the lung milieu is an ideal and familiar environment, thereby promoting the engraftment and differentiation of transplanted cells. Recent studies have shown the presence of multipotent ‘mesenchymal stem cells’ in the adult lung. The majority of these reports are, however, limited to animal models, and to date, there has been no report of a similar cell population in adult human lung parenchyma. Here, we show the identification of a population of primary human lung parenchyma (pHLP) mesenchymal stromal cells (MSCs) derived from intraoperative normal lung parenchyma biopsies. Surface and intracellular immunophenotyping by flow cytometry revealed that cultures do not contain alveolar type I epithelial cells or Clara cells, and are devoid of the following hematopoietic markers: CD34, CD45 and CXCR4. Cells show an expression pattern of surface antigens characteristic of MSCs, including CD73, CD166, CD105, CD90 and STRO-1. As per bone marrow MSCs, our pHLP cells have the ability to differentiate along the adipogenic, osteogenic and chondrogenic mesodermal lineages when cultured in the appropriate conditions. In addition, when placed in small airway growth media, pHLP cell cultures depict the expression of aquaporin 5 and Clara cell secretory protein, which is identified with that of alveolar type I epithelial cells and Clara cells, respectively, thereby exhibiting the capacity to potentially differentiate into airway epithelial cells. Further investigation of these resident cells may elucidate a therapeutic cell population capable of lung repair and/or regeneration.

A Novel Model for Post‐Transplant Obliterative Airway Disease Reveals Angiogenesis from the Pulmonary Circulation

We present a novel animal model for post‐transplant obliterative airway disease in which the donor trachea is implanted into the recipients lung parenchyma. Although this procedure is technically more challenging than the heterotopic model of implantation into a subcutaneous pouch, it has several important advantages some of which are the appropriate local environment and the possibility of local immunosuppressive therapy after transtracheal gene, cell or drug delivery. This model has revealed new insights into angiogenic potential of the pulmonary circulation.

Fluorescent microangiography (FMA): an improved tool to visualize the pulmonary microvasculature

Visualization of the complex lung microvasculature and resolution of its three-dimensional architecture remains a difficult experimental challenge. We present a novel fluorescent microscopy technique to visualize both the normal and diseased pulmonary microvasculature. Physiologically relevant pulmonary perfusion conditions were applied using a low-viscosity perfusate infused under continuous airway ventilation. Intensely fluorescent polystyrene microspheres, confined to the vascular space, were imaged through confocal optical sectioning of 200 μm-thick lung sections. We applied this technique to rat lungs and the markedly enhanced depth of field in projected images allowed us to follow vascular branching patterns in both normal lungs and lungs from animals with experimentally induced pulmonary arterial hypertension. In addition, this method allowed complementary immunostaining and identification of cellular components surrounding the blood vessels. Fluorescent microangiography is a widely applicable and quantitative tool for the study of vascular changes in animal models of pulmonary disease.

Recipient Treatment With Trimetazidine Improves Graft Function and Protects Energy Status After Lung Transplantation

Abstract Background Ischemia–reperfusion injury remains an important obstacle to successful lung transplantation. Trimetazidine is an anti-ischemic drug that restores the ability of ischemic cells to produce energy and reduces the generation of oxygen-derived free radicals. The aim of this study was to assess the protective effect of trimetazidine after prolonged ischemia in lung transplantation. Methods Rat single-lung transplantation was performed in 4 experimental groups (n = 5 each). In all groups, transplantation was performed after 18 hours of cold (4° C) ischemia. All donor lungs were flushed with low-potassium dextran–glucose (LPDG) solution that also contained 500 μg/liter prostaglandin estradiol (E 1 ). Groups studied included: Group I: flush solution was administered containing 10 −6 mol/liter trimetazidine (TMZ), neither donor nor recipient treatment given; Group II: donors were treated with 5 mg/kg intravenous TMZ 10 minutes prior to harvest, but the flush solution did not contain TMZ; Group III: recipients treated with 5 mg/kg intravenous TMZ 10 minutes before reperfusion, and flush solution contained 10 −6 mol/liter trimetazidine; Group IV: ischemic control group. After 2 hours of reperfusion, oxygenation was measured and lung tissue was frozen and assessed for adenosine triphosphate (ATP) content, myeloperoxidase (MPO) activity and thiobarbituric acid–reactive substances (TBARS). Peak airway pressure (PawP) was recorded throughout the reperfusion period. Results Group III showed significantly higher levels of ATP content (11.1 ± 5.01 pmol vs Group I, 3.36 ± 1.8 pmol, p = 0.008; vs Group II, 4.7 ± 1.9 pmol, p = 0.03; vs Group IV, 0.7 ± 0.2 pmol, p = 0.008), better oxygenation (442.5 ± 26.5 mm Hg, vs Group I, 161.06 ± 54.5 mm Hg; vs Group II, 266.02 ± 76.9 mm Hg; vs Group IV, 89.4 ± 14.7 mm Hg, p = 0.008) and reduced lipid peroxidation (TBARS) (0.15 ± 0.03 nmol/g; vs Group I, 1.04 ± 0.76 nmol/g; vs Group II, 0.69 ± 0.4 nmol/g; vs Group IV, 2.29 ± 0.4 nmol/g, p = 0.008). PawP and MPO activity were comparable in the 4 study groups. Conclusion Recipient treatment with TMZ provided significant protection of energy status, better oxygenation and reduced lipid peroxidation. Our data suggest that TMZ may be an important adjunct in the prevention of post-transplant lung ischemia–reperfusion injury.

Prevention of recurrent empyema after pneumonectomy for chronic infection

OBJECTIVES Pneumonectomy in chronic pulmonary infection with empyema is associated with a high mortality rate and an increased risk of recurrent empyema. The surgical resection is technically demanding, and successful management continues to be a challenge. METHODS We evaluated a concept which combines (pleuro-)pneumonectomy or completion pneumonectomy with surgical debridement of the pleural cavity and packing with povidine-iodine soaked dressings. The debridement and packing is repeated in the operating theater after 48 h until the chest cavity is macroscopically clean. Finally, the pleural space is obliterated with antibiotic solution. RESULTS Between February 1997 and October 2000, 11 patients (average age of 59 years, ranging from 25 to 84) with destroyed lung caused by tuberculosis (six), aspergilloma (two), bronchiectasis (one), esophago-pleural fistula (one) or broncho-pleural fistula after lobectomy for bronchial carcinoma (one) and ongoing chronic infection with acute empyema (ten) (25-2500 days between first and definitive therapy) were treated. Pleural culture findings showed Aspergillus in four, Mycobacterium in two, Enterococcus in two, Candida in one and Staphylococcus in one, respectively. The mean number of interventions was 2.9 (2-4). The chest was definitively closed in all patients within 1 week. The mean hospitalization time was 19 days (9-31 days). In the follow-up (10-54 months), there was no recurrence of empyema. One patient (84 years) died at day 31, due to sepsis. CONCLUSIONS Pneumonectomy combined with repeated surgical debridement and antimicrobial therapy enables the successful treatment of chronic pulmonary infection with empyema within a short time period.

Normal gas exchange after 30-h ischemia and treatment with phosphodiesterase inhibitor PDI747.

OBJECTIVE Phosphodiesterases (PDEs) negatively regulate the concentrations of cAMP and/or cGMP, which act as downstream second messengers to the prostaglandins. PDE type-4 (PDE4) is selective for cAMP and is found in high concentrations in endothelial, epithelial, and different blood cells. The aim of this study was to evaluate if PDI747, a novel selective inhibitor of PDE4, can restore pretransplant cAMP levels and thereby posttransplant organ function after prolonged cold ischemia. METHODS Left lung transplantation was performed in pigs (25-31 kg). Donor lungs were flushed with low potassium dextran glucose (LPDG) solution only (control, n=5)or, in addition with 1 micromol of PDI747 (PDI747, n=5) and stored for 30 h at 1 degrees C. PDI747 animals further received a bolus of PDI747 (0.3 mg/kg) 15 min prior to reperfusion and a continuous infusion (0.3 mg/kg per hour) during the 5 h after reperfusion. After occlusion of the right pulmonary arteries and the right main bronchus, hemodynamic and gas exchange parameters and extravascular lung water (EVLW) levels of the transplanted lung were assessed. RESULTS Two control animals died of severe lung edema leading to heart failure (control, n=3). One animal in the treatment group was excluded due to a patent ductus arteriosus (PDI747, n=4). Gas exchange at the end of the experiment was restored to normal levels in the PDI747 group (Pa, O(2) 47.6+/-11.2 kPa, Pa,CO(2) 6.4+/-1.8 kPa) but not in the control group (Pa, O(2) 7.7+/-2.9 kPa, Pa, CO(2) 11.9+/-3.0 kPa, P(PaO2)<0.0001, P(Pa, CO2)=0.06). Extravascular lung water (EVLW) was normal in the PDI747 group (8.5+/-1.1 ml/kg) and clearly elevated in the control group (16.2+/-5.6 ml/kg, P=0.007). Airway pressure in the PDI747 group was significantly lower than in the control group (7.8+/-0.5 cm H(2)O vs. 11.3+/-0.6 cm H(2)O, respectively, P<0.0001). The free radical mediated tissue injury measured by lipid peroxidation (TBARS) was significantly reduced (P=0.001) in the PDI747 group. CONCLUSIONS With the inhibition of PDE4 with PDI747 we achieved normal gas exchange, no posttransplant lung edema, normal airway pressures, and a reduced free radical injury after 30 h of cold ischemia.