Ryoichiro Doi

Ventilation-Based Decellularization System of the Lung

Abstract The demand for donated organs greatly exceeds the availability. Alternatives to organ donation, such as laboratory-engineered organs, are therefore being developed. One approach is to decellularize the organ and reseed it with selected cells, ideally from the organ recipient. Organ decellularization has typically been attempted by the administration of detergents into vessels such as the portal vein in the liver. However, in the case of the lung, the airway provides another potential administration route, because it has a wide contact area between cells and detergents in the tracheal tree and alveoli. In the present study, we introduce a novel ventilation-based decellularization system for the lung and compare its efficacy to ordinary decellularization systems administering detergent through the pulmonary artery. Rat lungs were decellularized using 500 mL of 3-[(3-cholamidopropyl) dimethylammonio]-1-Propanesulfonate (CHAPS) decellularization solution administrated through the pulmonary artery (vessel group) or through the trachea (airway group). The vessel group was infused CHAPS solution using a gravitational pressure head of 20 cmH2O. The airway group was infused with the detergent using negative pressure and positive end-expiratory pressure, for a volume 10cc with each inspiration in a bioreactor. Pathological and immunohistochemical findings indicated that components of the extracellular matrix (ECM), including proteoglycans, elastic fibers, fibronectin, and laminin, were more decreased in the airway group than in the vessel group. Western blot analysis showed that MHC class I antigen and β-actin were not detected in both decellularized groups. A collagen assay showed that collagen was 70% preserved in both groups compared to native lung. Glycosaminoglycan (GAG) and DNA assays showed that GAG and DNA contents were strongly diminished in both decellularized groups, but those contents were smaller in the airway group than in the vessel group. Accordingly, the alveolar wall was thinner on electron microscopy, and DNA remnants were not observed in the airway group. Infusion of red blood cells indicated that capillary walls were preserved without blood leakage in both groups. In conclusion, we describe a novel approach for decellularization through the airway that represents a more stringent method for both DNA and ECM removal, with capillary wall preservation.

Transplantation of bioengineered rat lungs recellularized with endothelial and adipose-derived stromal cells

Bioengineered lungs consisting of a decellularized lung scaffold that is repopulated with a patient’s own cells could provide desperately needed donor organs in the future. This approach has been tested in rats, and has been partially explored in porcine and human lungs. However, existing bioengineered lungs are fragile, in part because of their immature vascular structure. Herein, we report the application of adipose-derived stem/stromal cells (ASCs) for engineering the pulmonary vasculature in a decellularized rat lung scaffold. We found that pre-seeded ASCs differentiated into pericytes and stabilized the endothelial cell (EC) monolayer in nascent pulmonary vessels, thereby contributing to EC survival in the regenerated lungs. The ASC-mediated stabilization of the ECs clearly reduced vascular permeability and suppressed alveolar hemorrhage in an orthotopic transplant model for up to 3 h after extubation. Fibroblast growth factor 9, a mesenchyme-targeting growth factor, enhanced ASC differentiation into pericytes but overstimulated their proliferation, causing a partial obstruction of the vasculature in the regenerated lung. ASCs may therefore provide a promising cell source for vascular regeneration in bioengineered lungs, though additional work is needed to optimize the growth factor or hormone milieu for organ culture.

Pulmonary resection for lung cancer in nonagenarians: a report of five cases.

Lung cancer surgery in octogenarians has already become common in Japan due to the high proportion of elderly people and the progress in perioperative management, including anesthesia and video-assisted thoracic surgery. However, there have been few reports about the surgical treatment of lung cancer in nonagenarians. Five cases (mean age: 91.2 years; 3 men, 2 women) of non-small cell lung cancer in nonagenarians in whom pulmonary resection was performed are reported. All were adenocarcinomas; three were clinical stage IA, and two were IB. Three patients underwent lobectomy, and two underwent wedge resection. Mediastinal lymph node dissection was omitted in all patients. There were no fatal postoperative complications. During follow-up, one patient died due to fatal arrhythmia, and one patient who underwent wedge resection had local recurrence, but the others are alive in good condition. Pulmonary resection of lung cancer is possible for highly selected, nonagenarian patients without lymph node metastasis.

Sodium hydroxide based non-detergent decellularizing solution for rat lung

ABSTRACT Lung transplantation is the last option for the treatment of end stage chronic lung disorders. Because the shortage of donor lung organs represents the main hurdle, lung regeneration has been considered to overcome this hurdle. Recellularization of decellularized organ scaffold is a promising option for organ regeneration. Although detergents are ordinarily used for decellularization, other approaches are possible. Here we used high alkaline (pH12) sodium hydroxide (NaOH)-PBS solution without detergents for lung decellularization and compared the efficacy on DNA elimination and ECM preservation with detergent based decellularization solutions CHAPS and SDS. Immunohistochemical image analysis showed that cell components were removed by NaOH solution as well as other detergents. A Collagen and GAG assay showed that the collagen reduction of the NaOH group was comparable to that of the CHAPS and SDS groups. However, DNA reduction was more significant in the NaOH group than in other groups (p < 0.0001). The recellularization of HUVEC revealed cell attachment was not inferior to that of the SDS group. Ex vivo functional analysis showed 100% oxygen ventilation increased oxygen partial pressure as artificial hemoglobin vesicle-PBS solution passed through regenerated lungs in the SDS or NaOH group. It was concluded that the NaOH-PBS based decellularization solution was comparable to ordinal decellularizaton solutions and competitive in cost effectiveness and residues in the decellularized scaffold negligible, thus providing another potential option to detergent for future clinical usage.

Is early postoperative administration of pregabalin beneficial for patients with lung cancer?—randomized control trial

BACKGROUND Post-thoracotomy pain is an obstacle for lung-cancer patients even after introduction of less invasive surgical procedures. The aim of this prospective study was to evaluate if early postoperative administration of pregabalin is beneficial for patients with non-small cell lung cancer (NSCLC). METHODS We conducted a randomized open control trial. Patients with NSCLC were allocated randomly to epidural and nonsteroidal anti-inflammatory drug (NSAID) use for analgesia (control group) or pregabalin use (pregabalin group). Primary endpoint was the frequency of additional administration of a NSAID. Secondary endpoints were intensity of ongoing pain, frequency of neuropathic pain, and pain catastrophizing. RESULTS Seventy-two patients were registered and allocated. Thirty-four cases in the control group and 33 in the pregabalin group were assessed. Age, sex, body mass index (BMI), type of surgical procedure, type of lymph-node dissection, operation time, bleeding, duration of chest-tube insertion, and postoperative hospital stay between the two groups was not significantly different. Frequency of additional NSAID use between the control group (2±4 suppositories) and pregabalin group (2±3 suppositories) was not significantly different (P=0.62). Numeric Rating Scale (NRS) for the intensity of ongoing pain, frequency of neuropathic pain, and Pain Catastrophizing Scale (PCS) between each group were not significantly different at any time until 3 months after surgery. CONCLUSIONS Early postoperative administration of pregabalin is not beneficial for patients with NSCLC.