Jun Hirano

Insufficient autophagy in idiopathic pulmonary fibrosis

Autophagy, a process that helps maintain homeostatic balance between the synthesis, degradation, and recycling of organelles and proteins to meet metabolic demands, plays an important regulatory role in cellular senescence and differentiation. Here we examine the regulatory role of autophagy in idiopathic pulmonary fibrosis (IPF) pathogenesis. We test the hypothesis that epithelial cell senescence and myofibroblast differentiation are consequences of insufficient autophagy. Using biochemical evaluation of in vitro models, we find that autophagy inhibition is sufficient to induce acceleration of epithelial cell senescence and myofibroblast differentiation in lung fibroblasts. Immunohistochemical evaluation of human IPF biospecimens reveals that epithelial cells show increased cellular senescence, and both overlaying epithelial cells and fibroblasts in fibroblastic foci (FF) express both ubiquitinated proteins and p62. These findings suggest that insufficient autophagy is an underlying mechanism of both accelerated cellular senescence and myofibroblast differentiation in a cell-type-specific manner and is a promising clue for understanding the pathogenesis of IPF.

Insufficient autophagy promotes bronchial epithelial cell senescence in chronic obstructive pulmonary disease

Tobacco smoke-induced accelerated cell senescence has been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). Cell senescence is accompanied by the accumulation of damaged cellular components suggesting that in COPD, inhibition of autophagy may contribute to cell senescence. Here we look at whether autophagy contributes to cigarette smoke extract (CSE) - induced cell senescence of primary human bronchial epithelial cells (HBEC), and further evaluate p62 and ubiquitinated protein levels in lung homogenates from COPD patients. We demonstrate that CSE transiently induces activation of autophagy in HBEC, followed by accelerated cell senescence and concomitant accumulation of p62 and ubiquitinated proteins. Autophagy inhibition further enhanced accumulations of p62 and ubiquitinated proteins, resulting in increased senescence and senescence-associated secretory phenotype (SASP) with interleukin (IL)-8 secretion. Conversely, autophagy activation by Torin1, a mammalian target of rapamycin (mTOR inhibitor), suppressed accumulations of p62 and ubiquitinated proteins and inhibits cell senescence. Despite increased baseline activity, autophagy induction in response to CSE was significantly decreased in HBEC from COPD patients. Increased accumulations of p62 and ubiquitinated proteins were detected in lung homogenates from COPD patients. Insufficient autophagic clearance of damaged proteins, including ubiquitinated proteins, is involved in accelerated cell senescence in COPD, suggesting a novel protective role for autophagy in the tobacco smoke-induced senescence-associated lung disease, COPD.

Autophagy induction by SIRT6 through attenuation of insulin-like growth factor signaling is involved in the regulation of human bronchial epithelial cell senescence.

Cigarette smoke (CS)–induced cellular senescence has been implicated in the pathogenesis of chronic obstructive pulmonary disease, and SIRT6, a histone deacetylase, antagonizes this senescence, presumably through the attenuation of insulin-like growth factor (IGF)-Akt signaling. Autophagy controls cellular senescence by eliminating damaged cellular components and is negatively regulated by IGF-Akt signaling through the mammalian target of rapamycin (mTOR). SIRT1, a representative sirtuin family, has been demonstrated to activate autophagy, but a role for SIRT6 in autophagy activation has not been shown. Therefore, we sought to investigate the regulatory role for SIRT6 in autophagy activation during CS-induced cellular senescence. SIRT6 expression levels were modulated by cDNA and small interfering RNA transfection in human bronchial epithelial cells (HBECs). Senescence-associated β-galactosidase staining and Western blotting of p21 were performed to evaluate senescence. We demonstrated that SIRT6 expression levels were decreased in lung homogenates from chronic obstructive pulmonary disease patients, and SIRT6 expression levels correlated significantly with the percentage of forced expiratory volume in 1 s/forced vital capacity. CS extract (CSE) suppressed SIRT6 expression in HBECs. CSE-induced HBEC senescence was inhibited by SIRT6 overexpression, whereas SIRT6 knockdown and mutant SIRT6 (H133Y) without histone deacetylase activity enhanced HBEC senescence. SIRT6 overexpression induced autophagy via attenuation of IGF-Akt-mTOR signaling. Conversely, SIRT6 knockdown and overexpression of a mutant SIRT6 (H133Y) inhibited autophagy. Autophagy inhibition by knockdown of ATG5 and LC3B attenuated the antisenescent effect of SIRT6 overexpression. These results suggest that SIRT6 is involved in CSE-induced HBEC senescence via autophagy regulation, which can be attributed to attenuation of IGF-Akt-mTOR signaling.

Involvement of creatine kinase B in cigarette smoke-induced bronchial epithelial cell senescence.

Cigarette smoke induces damage to proteins and organelles by oxidative stress, resulting in accelerated epithelial cell senescence in the lung, which is implicated in chronic obstructive pulmonary disease (COPD) pathogenesis. Although the detailed molecular mechanisms are not fully understood, cellular energy status is one of the most crucial determinants for cell senescence. Creatine kinase (CK) is a constitutive enzyme, playing regulatory roles in energy homeostasis of cells. Among two isozymes, brain-type CK (CKB) is the predominant CK in lung tissue. In this study, we investigated the role of CKB in cigarette smoke extract (CSE)-induced cellular senescence in human bronchial epithelial cells (HBECs). Primary HBECs and Beas2B cells were used. Protein carbonylation was evaluated as a marker of oxidative protein damage. Cellular senescence was evaluated by senescence-associated β-galactosidase staining. CKB inhibition was examined by small interfering RNA and cyclocreatine. Secretion of IL-8, a hallmark of senescence-associated secretary phenotype, was measured by ELISA. CKB expression levels were reduced in HBECs from patients with COPD compared with that of HBECs from nonsmokers. CSE induced carbonylation of CKB and subsequently decreased CKB protein levels, which was reversed by a proteasome inhibitor. CKB inhibition alone induced cell senescence, and further enhanced CSE-induced cell senescence and IL-8 secretion. CSE-induced oxidation of CKB is a trigger for proteasomal degradation. Concomitant loss of enzymatic activity regulating energy homeostasis may lead to the acceleration of bronchial epithelial cell senescence, which is implicated in the pathogenesis of COPD.

Unilateral thoracoscopic subtotal thymectomy for the treatment of stage I and II thymoma

OBJECTIVE The purpose of this study was to determine the feasibility of thoracoscopic thymectomy for the treatment of Masaoka stage I and II thymoma. METHODS We evaluated the short-term outcomes of 40 patients undergoing surgery for Masaoka stage I and II thymomas without myasthenia gravis between July 2000 and July 2008. Of these, 22 patients underwent complete thymoma resection using unilateral thoracoscopic subtotal thymectomy (UTST group), and 18 patients underwent trans-sternal thymectomy (TST group). RESULTS Intra-operative blood loss amounts did not differ significantly between the UTST and TST groups (100.6 ml and 208.1 ml, respectively, p=0.0513). The duration of the postoperative hospital stay was significantly shortened in the UTST group (4.6 days vs 11.2 days, p<0.0001). No patient in the UTST group underwent conversion to open surgery. No severe surgical complications, such as bleeding due to injury to the left brachiocephalic vein, and no postoperative complications, were detected in this series. CONCLUSIONS These preliminary results suggest that thoracoscopic thymectomy for Masaoka stage I and II thymoma is technically feasible and safe, and it is less invasive for the patient. Nevertheless, this procedure requires further investigation in a large series with a longer follow-up.

Insulin-Dependent Phosphatidylinositol 3-Kinase/Akt and ERK Signaling Pathways Inhibit TLR3-Mediated Human Bronchial Epithelial Cell Apoptosis

TLR3, one of the TLRs involved in the recognition of infectious pathogens for innate and adaptive immunity, primarily recognizes viral-associated dsRNA. Recognition of dsRNA byproducts released from apoptotic and necrotic cells is a recently proposed mechanism for the amplification of toxicity, suggesting a pivotal participation of TLR3 in viral infection, as well as in lung diseases where apoptosis plays a critical role, such as asthma and chronic obstructive pulmonary disease. In addition to metabolic control, insulin signaling was postulated to be protective by inhibiting apoptosis. Therefore, we explored the role of insulin signaling in protecting against TLR3-mediated apoptosis of human bronchial epithelial cells. Significant TLR3-mediated apoptosis was induced by polyinosinic-polycytidylic acid, a dsRNA analog, via caspase-8–dependent mechanisms. However, insulin efficiently inhibited TLR3/ polyinosinic-polycytidylic acid-induced human bronchial epithelial cell apoptosis via PI3K/Akt and ERK pathways, at least in part, via upregulation of cellular FLIPs and through protein synthesis-independent mechanisms. These results indicate the significance of TLR3-mediated dsRNA-induced apoptosis in the pathogenesis of apoptosis-driven lung disease and provide evidence for a novel protective role of insulin.

Apoptosis inhibitor of macrophage (AIM) expression in alveolar macrophages in COPD

BackgroundMarked accumulation of alveolar macrophages (AM) conferred by apoptosis resistance has been implicated in pathogenesis of chronic obstructive pulmonary disease (COPD). Apoptosis inhibitor of macrophage (AIM), has been shown to be produced by mature tissue macrophages and AIM demonstrates anti-apoptotic property against multiple apoptosis-inducing stimuli. Accordingly, we attempt to determine if AIM is expressed in AM and whether AIM is involved in the regulation of apoptosis in the setting of cigarette smoke extract (CSE) exposure.MethodsImmunohistochemical evaluations of AIM were performed. Immunostaining was assessed by counting total and positively staining AM numbers in each case (n = 5 in control, n = 5 in non-COPD smoker, n = 5 in COPD). AM were isolated from bronchoalveolar lavage fluid (BALF). The changes of AIM expression levels in response to CSE exposure in AM were evaluated. Knock-down of anti-apoptotic Bcl-xL was mediated by siRNA transfection. U937 monocyte-macrophage cell line was used to explore the anti-apoptotic properties of AIM.ResultsThe numbers of AM and AIM-positive AM were significantly increased in COPD lungs. AIM expression was demonstrated at both mRNA and protein levels in isolated AM, which was enhanced in response to CSE exposure. AIM significantly increased Bcl-xL expression levels in AM and Bcl-xL was involved in a part of anti-apoptotic mechanisms of AIM in U937 cells in the setting of CSE exposure.ConclusionsThese results suggest that AIM expression in association with cigarette smoking may be involved in accumulation of AM in COPD.

Anomalous Systemic Arterial Supply to Separate Lingular and Basal Segments of the Lung: An Anatomic Consideration

A 22-year-old man was referred for hemoptysis and general fatigue after exercise. Arteriography demonstrated an anomalous artery arising from the descending aorta supplying the lingular and all of the basal segments of the left lung. The feeding areas of the pulmonary and anomalous arteries were mutually exclusive. He underwent division of the anomalous artery and combined resection of the diseased segments. The upper division of the upper lobe and the superior segment of the lower lobe were spared. His symptoms were greatly improved postoperatively. The preoperative anatomic evaluation of anomalous vessels is crucial in surgical management.

Study on surgical treatment for lung cancer associated with giant bullous disease

Five patients of primary lung cancer with giant bullous disease underwent surgery from April 1985 to December 1995. All patients were male and heavy smokers, and the median age was 50 years. The location of the tumor was in the right upper lobe in four patients and in the left upper lobe in the other. Three patients were treated by lobectomy and two by sleeve lobectomy. Histological examination showed large cell carcinoma in four patients and poorly-differentiated adenocarcinoma in the other. The pathological stage was I in three. IIIA in one, and IV in the other. Two of three in stage I have survived for more than 6 years postopertively without recurrence, and the other died of brain metastasis. The stage IIIA case and the IV case died 3 years and one year postoperatively, respectively. The clinical features of lung cancer associated with giant bullous disease was discussed by reviewing 33 patients reported in Japan, including our patients. In 13 patients, lung cancer and bullous disease were diagnosed simultaneously (group A), and in 20 patients, bullous disease were diagnosed prior to the appearance of an abnormal shadow due to lung cancer (group B). The patients in group B had a tendency to be diagnosed at an earlier stage of lung cancer than the patients in group A. In the patients of stage I, the 5-year survival rate was 78.6%, however, in the patients of more than stage IIIA, 3-year survival rate was 26.5% and the 5-year survival rate was 0%. Significant differences in the survival curves were demonstrated between the cases with stage I and the cases with more than stage IIIA. In conclusion, in order to improve the prognosis of lung cancer with giant bullous disease, it is considered to be important to detect giant bulla prior to lung cancer, and when a case of bullous disease is found, periodical follow-up must be done to find early stage lung cancer.

Thoracoscopic resection of a mediastinal venous hemangioma: Report of a case

Mediastinal hemangioma is a rare tumor, accounting for 0.5% or fewer of all mediastinal tumors in most reports. We herein report a case of thoracoscopic resection of venous hemangioma in the middle mediastinum. A 48-year-old man was referred to our hospital for dysphagia. A chest computed tomography scan showed the mass to have a smooth surface and heterogeneous contents. Magnetic resonance imaging demonstrated the morphology of the vascular tumor to be a hyperintense mass on a T2-weighted image. Using thoracoscopic surgery, the tumor was completely extirpated and confirmed histologically to be a venous hemangioma. In this case, thoracoscopic surgery provided a satisfactory view and facilitated correct handling for the mediastinal venous hemangioma.