Barbara C. Cahill

NOX4 mediates hypoxia-induced proliferation of human pulmonary artery smooth muscle cells: the role of autocrine production of transforming growth factor-β1 and insulin-like growth factor binding protein-3

Persistent hypoxia can cause pulmonary arterial hypertension that may be associated with significant remodeling of the pulmonary arteries, including smooth muscle cell proliferation and hypertrophy. We previously demonstrated that the NADPH oxidase homolog NOX4 mediates human pulmonary artery smooth muscle cell (HPASMC) proliferation by transforming growth factor-beta1 (TGF-beta1). We now show that hypoxia increases HPASMC proliferation in vitro, accompanied by increased reactive oxygen species generation and NOX4 gene expression, and is inhibited by antioxidants, the flavoenzyme inhibitor diphenyleneiodonium (DPI), and NOX4 gene silencing. HPASMC proliferation and NOX4 expression are also observed when media from hypoxic HPASMC are added to HPASMC grown in normoxic conditions, suggesting autocrine stimulation. TGF-beta1 and insulin-like growth factor binding protein-3 (IGFBP-3) are both increased in the media of hypoxic HPASMC, and increased IGFBP-3 gene expression is noted in hypoxic HPASMC. Treatment with anti-TGF-beta1 antibody attenuates NOX4 and IGFBP-3 gene expression, accumulation of IGFBP-3 protein in media, and proliferation. Inhibition of IGFBP-3 expression with small interfering RNA (siRNA) decreases NOX4 gene expression and hypoxic proliferation. Conversely, NOX4 silencing does not decrease hypoxic IGFBP-3 gene expression or secreted protein. Smad inhibition does not but the phosphatidylinositol 3-kinase (PI3K) signaling pathway inhibitor LY-294002 does inhibit NOX4 and IGFBP-3 gene expression, IGFBP-3 secretion, and cellular proliferation resulting from hypoxia. Immunoblots from hypoxic HPASMC reveal increased TGF-beta1-mediated phosphorylation of the serine/threonine kinase (Akt), consistent with hypoxia-induced activation of PI3K/Akt signaling pathways to promote proliferation. We conclude that hypoxic HPASMC produce TGF-beta1 that acts in an autocrine fashion to induce IGFBP-3 through PI3K/Akt. IGFBP-3 increases NOX4 gene expression, resulting in HPASMC proliferation. These observations add to our understanding hypoxic pulmonary vascular remodeling.

EIF2AK4 Mutations in Pulmonary Capillary Hemangiomatosis

BACKGROUND Pulmonary capillary hemangiomatosis (PCH) is a rare disease of capillary proliferation of unknown cause and with a high mortality. Families with multiple affected individuals with PCH suggest a heritable cause although the genetic etiology remains unknown. METHODS We used exome sequencing to identify a candidate gene for PCH in a family with two affected brothers. We then screened 11 unrelated patients with familial (n = 1) or sporadic (n = 10) PCH for mutations. RESULTS Using exome sequencing, we identified compound mutations in eukaryotic translation initiation factor 2 α kinase 4 (EIF2AK4) (formerly known as GCN2) in both affected brothers. Both parents and an unaffected sister were heterozygous carriers. In addition, we identified two EIF2AK4 mutations in each of two of 10 unrelated individuals with sporadic PCH. EIF2AK4 belongs to a family of kinases that regulate angiogenesis in response to cellular stress. CONCLUSIONS Mutations in EIF2AK4 are likely to cause autosomal-recessive PCH in familial and some nonfamilial cases.

Early experience with sirolimus in lung transplant recipients with chronic allograft rejection

BACKGROUND Chronic lung allograft rejection, commonly manifest as obliterative bronchiolitis (OB/BOS), hinders long-term survival after lung transplantation (LT). OB/BOS is traditionally treated with augmented immunosuppression and results in short-term stabilization in pulmonary function for most patients. However, peribronchiolar fibroproliferation and airway obstruction usually recur despite initial improvements seen with increases in immunosuppression. In this observational, uncontrolled study, the effect of sirolimus, a novel immunosuppressant with anti-proliferative activity, was assessed in LT patients with OB/BOS. METHODS Between June 1999 to November 2000, LT recipients with newly diagnosed or progressive OB/BOS received sirolimus in combination with a calcineurin inhibitor (CI) and prednisone. Pulmonary function, laboratory data and adverse effects were monitored for the first 24 weeks of therapy. RESULTS Sirolimus was utilized in 12 LT recipients with OB/BOS. After drug initiation, 58% of patients required a reduction in CI dose to maintain appropriate CI trough concentrations. Despite CI dose reduction, serum creatinine rose in 75% of patients. Unexpected adverse effects included anemia of chronic disease (100%), edema (50%) and malignancy (17%). For the group, the rate of change in FEV(1) and FEF(25%-75%) was unchanged with sirolimus, but individual responses varied. CONCLUSIONS For the group, the decline in pulmonary function was not affected by the addition of sirolimus. However, among individuals with rapidly declining pulmonary mechanics, sirolimus resulted in stabilization or improvement in pulmonary function. Significant adverse effects resulted from combination sirolimus plus CI therapy. Until optimal dosing strategies and a more complete adverse effect profile are established, combination therapy should be utilized cautiously in these patients.

Nocardia veterana as a Pathogen in North American Patients

ABSTRACT The molecular methodologies used in our laboratories have allowed us to define a group of Nocardia isolates from clinical samples which resemble the type strain of Nocardia veterana. Three patient isolates and the type strain of N. veterana gave identical and distinctive restriction fragment length polymorphisms (RFLPs) for an amplified portion of the 16S rRNA gene. These three isolates and the N. veterana type strain also gave identical RFLPs for an amplified portion of the 65-kDa heat shock protein gene, but this pattern was identical to that obtained for the Nocardia nova type strain. Sequence analysis of both a 1,359-bp region of the 16S rRNA gene and a 441-bp region of the heat shock protein gene of the patient isolates showed 100% identities with the same regions of the N. veterana type strain. DNA-DNA hybridization of the DNA of one of the patient isolates with the DNA of the N. veterana type strain showed a relative binding ratio of 82%, with 0% divergence, confirming that the isolate was N. veterana. Biochemical and susceptibility testing showed no significant differences among the patient isolates and the N. veterana type strain. Significantly, the results of antimicrobial susceptibility testing obtained for our isolates were similar to those obtained for N. nova, indicating that susceptibility testing alone cannot discriminate between these species. We present two case studies which show that N. veterana is a causative agent of pulmonary disease in immunocompromised patients residing in North America. We also describe difficulties encountered in using 16S rRNA gene sequences alone for discrimination of N. veterana from the related species Nocardia africana and N. nova because of the very high degree of 16S rRNA gene similarity among them.

Prevention of bone loss and fracture after lung transplantation: A pilot study

Background. Osteoporotic fracture is a significant source of morbidity after lung transplantation. Therapies to prevent posttransplant fracture are largely untested among lung transplant recipients. Methods. In this prospective uncontrolled study, lung transplant referrals were assessed for bone health with metabolic, radiographic, and bone mineral density measurements. Transplant recipients were treated with an antiresorptive regimen that included a bisphosphonate starting before or after transplantation. One year after transplantation, the fracture rate and bone density of patients in each group were reassessed and compared to historical controls. Between January 1996 and August 1999, 45/50 (90%) lung transplant referrals underwent bone health assessment. Transplant candidates received calcium, vitamin D, and hormone replacement therapy as indicated for hypogonadism. After July 1998, bisphosphonate therapy was added for candidates with osteopenia or osteoporosis (T score <1). After transplantation, all patients received 90 mg of pamidronate i.v. every 12 weeks, regardless of pretransplant bone density. Radiologic evaluation was performed for clinical suspicion of fracture. Bone density was remeasured 1 year after transplantation. Results. Most transplant referrals suffered from osteopenia or osteoporosis, and 29% of transplant referrals had prevalent vertebral compression fractures. Hypogonadism was untreated in 50% of men and 20% of women, and 15% of patients had hypovitaminosis D. Of the 21 patients assessed 1 year after transplantation, new fractures occurred in 4% of these patients. Lateral lumbar spine and hip bone density remained stable or improved in 65% and 86% of patients, respectively. Most of those who lost bone density had started bisphosphonate therapy after transplantation. Conclusions. Antiresorptive therapy with a bisphosphonate decreases the fracture rate and preserves bone mass 1 year after lung transplantation. In end-stage lung disease patients with osteopenia or osteoporosis, bisphosphonate therapy should be initiated before transplant surgery is contemplated.

Lung transplantation for cystic fibrosis.

Purpose of review To examine recent publications on lung transplantation for cystic fibrosis for changes in surgical techniques, selection criteria of patients, and impact on quality of life. Recent findings Recent evidence focuses on cystic fibrosis patient subsets enabling better decisions about listing for lung transplantation as a therapeutic option. There is information about Burkholderia cepacia infection, ventilator dependence, young age, and arthropathy. In the US, the United Network for Organ Sharing has addressed perceived inequities in organ distribution by allocating organs by illness severity rather than time on the waiting list. A Lung Allocation Score ranks severity for patients 12 years of age and older for transplantation based on variables including lung function, oxygen and ventilatory needs, diabetes, weight and physical performance. Some recently studied important variables that influence survival in cystic fibrosis and after lung transplantation, including airway infections, pancreatic exocrine function and acute exacerbations, are not included in the Lung Allocation Score. Few publications have examined quality of life after transplantation, and a definitive work has yet to appear. Summary New information has refined decision-making about lung transplantation for patients with cystic fibrosis. We examine recent findings and make recommendations for patients, families and medical providers.

Selection of patients with cystic fibrosis for lung transplantation.

Lung transplantation is the most aggressive therapy available for end-stage lung disease from cystic fibrosis (CF). A new predictive survival model of CF uses demographic, FEV1, nutritional, microbiologic, and acute exacerbation data to produce precise estimates of 5-year survival. The model improves the ability to select patients most likely to have survival benefit from transplantation. We discuss potential application of the survival model to four distinct groups of patients with CF: (1) candidates for cadaveric transplantation, (2) potential living donor recipients, (3) patients infected with multiply-resistant organisms such as Burkholderia cepacia, and (4) patients critically ill and dependent on mechanical ventilation. Measuring the impact of transplantation on quality of life remains a difficult task, and further studies are needed to determine whether lung-transplantation-derived survival benefit implies quality-of-life benefit. However, judicious use of the survival model to select patients for transplantation is likely to improve survival outcomes.

Tracheobronchial metal stents: Effects of covering a bronchial ostium in pigs

RATIONALE AND OBJECTIVES The purpose of this study was to examine the effects of placing a metal stent across a bronchial orifice. MATERIALS AND METHODS Nine pigs were used as test subjects, because the right upper lobe bronchus comes directly off the trachea in these animals. One of three types of metal stents was placed into the trachea of each pig and covered the orifice of the right upper lobe bronchus. Follow-up studies were performed at 1 and 3 months to evaluate the right upper lobe for signs of bronchial obstruction, infection, and atelectasis. The animals were sacrificed at 3 months to study the histopathologic changes of the trachea and lungs. RESULTS Two upper lobe bronchi remained patent; seven were obstructed by granulation tissue or plugs of mucus and inflammatory cells. Right upper lobe infiltration and atelectasis were seen in eight animals. Interestingly, radiographic opacities were also common in other lung segments. There was a tendency toward fewer and less extensive lung opacities at 3 months compared with that at 1 month. At histopathologic examination, areas of both acute and chronic pneumonia were found in the right upper lobe of all animals. The segment of trachea covered by the stent was lined with a thin layer of granulation tissue containing neutrophils, monocytes, and lymphocytes. The stent luminal surface was covered with columnar, cuboidal, and stratified squamous epithelium. Tracheal stenosis was seen in three animals because of excessive granulation tissue in two and a collapsed stent in one. CONCLUSION Placement of metal stents in pig trachea covering the orifice of the right upper lobe bronchus resulted in retention of secretions and secondary infection in the right upper lobe and other distant lung segments.

Lung transplantation following coronary artery bypass surgery—improved outcomes following single-lung transplant

BACKGROUND Prior coronary artery bypass grafting (CABG) has been a contraindication to lung transplantation (LTx) because of disease severity and technical considerations. Although patients increasingly are being referred for and receiving LTx, whether it should remain a contraindication is unknown. We sought to define the prevalence of LTx after CABG and determine the effect on outcomes. METHODS The United Network for Organ Sharing Standard Transplant Analysis and Research data set was queried during the period 2004-2013 for adult LTx patients, as prior CABG became a mandatory reporting field in 2004. The primary end-points were 30-day and 1-, 3-, and 5-year survivals. RESULTS The study cohort included 14,791 patients, of whom 292 patients had previously undergone CABG (single left, n = 68; single right, n = 181; bilateral, n = 43), representing 2% of all transplants. For the entire cohort, 30-day survival was 97%, and survival at 1, 3, and 5 years was 88%, 79%, and 74%. CABG was a predictor of mortality at all time points, with hazard ratios ranging from 1.97 (confidence interval, 1.23-3.16; p < 0.01) at 30 days to 1.38 (confidence interval, 1.12-1.69; p < 0.01) at 5 years. When stratified by type of transplant, CABG strongly predicted mortality at all time points for patients receiving bilateral, but not single, transplants. CONCLUSIONS Although LTx after CABG is uncommon, it is increasingly performed in the current era. Single right LTx is the most common procedure performed in patients with prior CABG. CABG before LTx is an independent predictor of mortality at all time points and is driven by increased mortality in patients receiving bilateral LTx.

Clinical course and spectrum of intensive care unit patients reactivating herpes simplex-1 virus: A retrospective analysis

Background: Herpes simplex-1 virus (HSV-1) reactivation in the respiratory tract is common in intensive care unit (ICU) patients. However, susceptible ICU populations are poorly defined. Clinical recognition of HSV infection of the respiratory tract is difficult and the impact of such reactivation is not understood. Materials and Methods: A retrospective analysis of HSV-1 positive patients encountered over a 5-year period at a multispecialty ICU was carried out. HSV-1 was identified in respiratory secretions using a qualitative polymerase chain reaction (PCR) technique. Patient charts were reviewed for clinical features that would typify HSV-1 respiratory involvement, and the morbidity and mortality risks found with HSV-1 respiratory involvement. Results: A review of 48 HSV-1 positive ICU patients showed that patients reactivating HSV in the respiratory tract fell into one of the three categories: (1) septic elderly patients with and without ARDS, (2) immunosuppressed patients, especially those receiving high-dose steroids, and (3) post-thoracotomy patients. Abnormalities suggestive of HSV-1 reactivation in the respiratory tract included, haemorrhagic or excessive respiratory secretions, concomitant orofacial herpes (42%), and bronchoscopic abnormalities (hemorrhagic ulcers and mucosal friability) (83%). Twenty eight percent of the HSV-1 infected patients experienced postextubation stridor. HSV-1 reactivation was associated with extended ventilator stays, significant mortality (42%), and ventilator-associated pneumonias (52%). Conclusions: Identification of susceptible populations and definition of clinical features of HSV-1 related respiratory disease can enable diagnosis of HSV-1 infection in ICU patients. Although detection by a PCR technique can rapidly diagnose HSV-1 reactivation, prospective studies are required to clarify HSV disease versus mere shedding, and understand the impact of HSV-1 reactivation in hospitalized patients.