Intraprocedural computed tomography-guided navigation with ventilatory strategy for atelectasis (ICNVA): a modified electromagnetic navigation bronchoscopy.

Background: Computed tomography (CT)-body divergence limits the accuracy of electromagnetic navigation bronchoscopy (ENB) in peripheral lung lesions diagnosis. We developed intraprocedural CT-guided navigation with ventilatory strategy for atelectasis (ICNVA) ENB for patients with peripheral lung lesions. Methods: Retrospective observational study in which ten consecutive patients with pulmonary lesions (without bronchial direct connection) underwent ICNVA-ENB was conducted. During ICNVA-ENB, intraoperative CT data were used for ENB path planning, and a new ventilation strategy were employed to help maintain the pulmonary region in a static and inflation state which reduce CT to body divergence. We collected three sets of CT data: preENB CT, post-anesthesia intubation CT, and postENB CT. To evaluate the accuracy of ICNVA-ENB, we measured the distance between the ENB probe and the actual lesion location, but also recorded the results of rapid on-site evaluation (ROSE), and postoperative pathology. To evaluate the impact of CT-body divergence induced by atelectasis, we calculated the mutual position distance of target lesions in preENB CT, post-anesthesia intubation CT and postENB CT. Furthermore, ENB operation time and operative complications were recorded. Results: Our analysis revealed that the distance between the navigation probe with the actual location of lesion center was 4-10 (5.90±1.73) mm. The ROSE results were consistent with the postoperative pathological diagnosis in 9 out of 10 patients (90%). The ICNVA-ENB atelectasis CT-body divergence was smaller than traditional ENB (12.10±3.67 vs. 6.60±2.59 mm, P<0.01). The ENB operation time was 20-53 (29.30±10.14) minutes and one patient developed slight intrapulmonary hemorrhage. Conclusions: ICNVA-ENB can reduce the CT-body divergence and appears to be safe and accurate for patients with peripheral lung lesions.

Single-center experience of transitioning from video-assisted laparoscopic to robotic Heller myotomy with Dor fundoplication for esophageal motility disorders.

BACKGROUND: Video-assisted laparoscopic Heller myotomy (LHM) has become the standard treatment option for achalasia. While robotic surgery offering some specific advantages such as better three-dimensional (3D) stereoscopic vision, hand-eye consistency, and flexibility and stability with the endowrist is expected to be shorter in learning curve than that of LHM for surgeons who are proficient in LHM. The aim of this study was to describe a single surgeon's experience related to the transition from video-assisted laparoscopic to robotic Heller myotomy with Dor fundoplication. METHODS: We conducted a retrospective observational study based on the recorded data of the first 66 Heller myotomy performed with laparoscopic Heller myotomy with Dor fundoplication (LHMD, 26 cases) and with the robotic Heller myotomy with Dor fundoplication (RHMD, 40 cases) by the same surgeon in Department of Thoracic Surgery of The First Affiliated Hospital of Nanchang University in China. The operation time and intraoperative blood loss were analyzed using the cumulative sum (CUSUM) method. Corresponding statistical tests were used to compare outcomes of both serials of cases. RESULTS: The median operation time was shorter in the RHMD group compared to the LHMD group (130 [IQR 123-141] minutes vs. 163 [IQR 153-169]) minutes, p < 0.001). In the RHMD group, one patient (2.5%) experienced mucosal perforation, whereas, in the LHMD group, the incidence of this complication was significantly higher at 19.2% (5 patients) (p = 0.031). Based on cumulative sum analyses, operation time decreased starting with case 20 in the LHMD group and with case 18 in the RHMD group. Intraoperative blood loss tended to decline starting with case 19 in the LHMD group and with case 16 in the RHMD group. CONCLUSIONS: Both RHMD and LHMD are effective surgical procedures for symptom relief of achalasia patients. RHMD demonstrates superior outcomes in terms of operation time and mucosal perforation during surgery compared to LHMD. Proficiency with RHMD can be achieved after approximately 16-18 cases, while that of LHMD can be obtained after around 19-20 cases.

microRNA-145 Inhibition Upregulates SIRT1 and Attenuates Autophagy in a Mouse Model of Lung Ischemia/Reperfusion Injury via NF-κB-dependent Beclin 1.

BACKGROUND: MicroRNA-145 (miR-145) has been shown to play a critical role in ischemia/reperfusion (I/R) injury; however, the expression and function of miR-145 in lung I/R injury have not been reported yet. This study aimed to elucidate the potential effects of miR-145 in lung I/R injury. METHODS: Lung I/R mice models and hypoxia/reoxygenation (H/R) pulmonary microvascular endothelial cell models were established. The expression of miR-145 and sirtuin 1 (SIRT1) was measured with reverse transcription-quantitative polymerase chain reaction and Western blot analysis in mouse lung tissue and cells. Artificial modulation of miR-145 and SIRT1 (downregulation) was done in I/R mice and H/R cells. Additionally, Pao2/FiO2 ratio, wet weight-to-dry weight ratio, and cell apoptosis in mouse lung tissues were determined by blood gas analyzer, electronic balance, and deoxyuridine triphosphate-biotin nick end-labeling assay, respectively. Autophagy marker Beclin 1 and LC3 expression, NF-κB acetylation levels, and autophagy bodies were detected in cell H/R and mouse I/R models by Western blot analysis. pulmonary microvascular endothelial cell apoptosis was detected with flow cytometry. RESULTS: miR-145 was abundantly expressed in the lung tissue of mice and PMVECs following I/R injury. In addition, miR-145 directly targeted SIRT1, which led to significantly decreased Pao2/FiO2 ratio and increased wet weight-to-dry weight ratio, elevated acetylation levels and transcriptional activity of NF-κB, upregulated expressions of tumor necrosis factor-α, interleukins-6, and Beclin 1, autophagy bodies, cell apoptosis, as well as LC3-II/LC3I ratio. CONCLUSIONS: In summary, miR-145 enhances autophagy and aggravates lung I/R injury by promoting NF-κB transcriptional activity via SIRT1 expression.

Effects of prolonged cold-ischemia on autophagy in the graft lung in a rat orthotopic lung transplantation model.

INTRODUCTION: Ischemia-reperfusion (I/R) injury causes present challenges in the field of graft transplantation which is also a major contributor to early graft dysfunction or failure after organ transplantation. The study focuses on the effects of prolonged cold-ischemia (CI) on the autophagic activity in the graft lung in a rat orthotopic lung transplantation model. MATERIAL AND METHODS: Donor lungs were preserved under CI conditions for different periods. An orthotopic lung transplantation model was developed, and the lung tissues from donor lungs subjected to CI preservation and reperfusion were harvested. We evaluated the effects of different CI periods on autophagy, reactive oxygen species (ROS) and glucose consumption. Additionally, the mechanism by which prolonged CI affected autophagy was investigated through determination of the molecules related to the mTOR pathway after treatment with 3-Methyladenine (3-MA), rapamycin and an adenosine triphosphate (ATP) synthase inhibitor oligomycin (OM). RESULTS: Prolonged CI led to increased activities of key glycolytic enzymes, glucose consumption and lactic acid production. Autophagy, ROS and glucose consumption were induced in the graft lung after I/R, which reached peak levels after 6 h and was gradually decreased. Most importantly, the perfusion treatment of 3-MA or OM decreased ROS level and autophagy, but increased the extent of mTOR phosphorylation, while the perfusion treatment of rapamycin induced ROS and autophagy. CONCLUSION: Taken together, autophagy mediated by a prolonged CI preservation affects the glucose consumption and ROS production in the graft lung via the mTOR signaling pathway.

microRNA-223 promotes autophagy to aggravate lung ischemia-reperfusion injury by inhibiting the expression of transcription factor HIF2α.

Lung ischemia-reperfusion (I/R) injury severely endangers human health, and recent studies have suggested that certain microRNAs (miRNAs) play important roles in this pathological phenomenon. The current study aimed to ascertain the ability of miR-223 to influence lung I/R injury by targeting hypoxia-inducible factor-2α (HIF2α). First, mouse models of lung I/R injury were established: during surgical procedures, pulmonary arteries and veins and unilateral pulmonary portal vessels were blocked and resuming bilateral pulmonary ventilation, followed by restoration of bipulmonary ventilation. In addition, a lung I/R injury cell model was constructed by exposure to hypoxic reoxygenation (H/R) in mouse pulmonary microvascular endothelial cells (PMVECs). Expression of miR-223, HIF2α, and ß-catenin in tissues or cells was determined by RT-qPCR and Western blot analysis. Correlation between miR-223 and HIF2α was analyzed by dual luciferase reporter gene assay. Furthermore, lung tissue injury and mouse PMVEC apoptosis was evaluated by hematoxylin and eosin (H&E), TUNEL staining, and flow cytometry. Autophagosomes in cells were detected by light chain 3 immunofluorescence assay. miR-223 was expressed at a high level while HIF2α/ß-catenin was downregulated in tissues and cells with lung I/R injury. Furthermore, miR-223 targeted and repressed HIF2α expression to downregulate ß-catenin expression. The miR-223/HIF2α/ß-catenin axis aggravated H/R injury in mouse PMVECs and lung I/R injury in mice by enhancing autophagy. Taken together, miR-223 inhibits HIF2α to repress ß-catenin, thus contributing to autophagy to complicate lung I/R injury. These findings provide a promising therapeutic target for treating lung I/R injury.

Notch1 protects against myocardial ischaemia-reperfusion injury via regulating mitochondrial fusion and function.

Mitochondrial fusion and fission dynamic are critical to the myocardial protection against ischaemia-reperfusion injury. Notch1 signalling plays an important role in heart development, maturation and repair. However, the role of Notch1 in the myocardial mitochondrial fusion and fission dynamic remains elusive. Here, we isolated myocardial cells from rats and established myocardial ischaemia-reperfusion injury (IRI) model. We modulated Notch1, MFN1 and DRP1 expression levels in myocardial cells via infection with recombinant adenoviruses. The results showed that Notch1 improves the cell viability and mitochondrial fusion in myocardiocytes exposed to IRI. These improvements were dependent on the regulation of MFN1 and DRP1. On the mechanism, we found that MNF1 is transcriptionally activated by RBP-Jk in myocardiocytes. Notch1 also improves the mitochondrial membrane potential in myocardiocytes exposed to IRI. Moreover, we further confirmed the protection of the Notch1-MFN1/Drp1 axis on the post-ischaemic recovery of myocardial performance is associated with the preservation of the mitochondrial structure. In conclusion, this study presented a detailed mechanism by which Notch1 signalling improves mitochondrial fusion during myocardial protection.

MicroRNA-497-5p negatively regulates the proliferation and cisplatin resistance of non-small cell lung cancer cells by targeting YAP1 and TEAD1.

BACKGROUND: MicroRNAs (miRNAs) are crucial regulators in the pathological processes and drug resistance of lung cancer. In this study, we investigated the role of miR-497-5p in modulating the function of non-small cell lung cancer (NSCLC). METHODS: MiR-497-5p expression in lung cancer tissues and cells was evaluated by qRT-PCR. Cell proliferation was evaluated by CCK-8 assay and colony-formation assay. Cell cycle and cell apoptosis were detected by flow cytometry. The effect of miR-497-5p on the expression of Yes-associated protein 1 (YAP1) and TEA domain family member 1 (TEAD1) was analyzed by qRT-PCR, Western blot and luciferase activity assay. RESULTS: The expression of miR-497-5p was significantly downregulated in lung cancer tissues and cells compared with paired normal tissues and cells. Overexpression of miR-497-5p induced growth retardation and apoptosis of A549 lung cancer cells. Mechanistically, YAP1 and TEAD1 were targeted and downregulated by miR-497-5p. Finally, we found that miR-497-5p increased cisplatin chemosensitivity in A549 cells. CONCLUSIONS: MiR-497-5p suppresses cell proliferation and resistance to cisplatin in NSCLC by downregulating the expression of YAP1 and TEAD1.

Methylglyoxal: A newly detected and potentially harmful metabolite in the blood of ketotic dairy cows.

Ketosis causes serious economic losses for the modern dairy industry because it is a highly prevalent metabolic disease among cows in high-producing herds during the transition period. Due to some striking similarities between diabetes in humans and ketosis in dairy cows, there is potential for the use of methylglyoxal (MGO)-commonly used in human diabetics-as a biomarker in dairy cattle. However, currently no data are available about the presence of MGO in the serum of dairy cattle or about the characteristics of its production or its potential contribution in the pathogenesis of ketosis. To determine the potential origin and pathway of formation of MGO, cows in different metabolic conditions [i.e., non-subclinically ketotic dairy cows in early lactation (n = 7), subclinically ketotic dairy cows in early lactation (n = 8), overconditioned dry cows (BCS >4.25, n = 6), and nonlactating heifers (n = 6)] were selected. Serum MGO concentrations were determined and correlated with indicators of the glucose and lipid metabolism and with haptoglobin (Hp) as an inflammatory marker. The serum MGO concentrations in subclinically ketotic cows (712.60 ± 278.77 nmol/L) were significantly greater than in nonlactating heifers (113.35 ± 38.90 nmol/L), overconditioned dry cows (259.71 ± 117.97 nmol/L), and non-subclinically ketotic cows (347.83 ± 63.56 nmol/L). In serum of lactating cows, concentrations of glucose and fructosamine were lower than in heifers and were negatively correlated with MGO concentrations. Even so, concentrations of metabolic and inflammatory markers such as dihydroxyacetone phosphate, nonesterified fatty acids, ß-hydroxybutyrate, acetone, and Hp were remarkably higher in subclinically ketotic cows compared with nonlactating heifers; these metabolites were also positively correlated with MGO. In human diabetics elevated MGO concentrations are stated to originate from both hyperglycemia and the enhanced lipid metabolism, whereas higher MGO concentrations in subclinically ketotic cows were not associated with hyperglycemia. Therefore, our data suggest MGO in dairy cows to be a metabolite produced from the metabolization of acetone within the lipid metabolization pathway and from the metabolization of dihydroxyacetone phosphate. Furthermore, the highly positive correlation between MGO and Hp suggests that this reactive compound might be involved in the proinflammatory state of subclinical ketosis in dairy cows. However, more research is needed to determine the potential use of MGO as a biomarker for metabolic failure in dairy cows.

Inhibition of Methylglyoxal-Induced Histone H1 Nε-Carboxymethyllysine Formation by (+)-Catechin.

Reactive dicarbonyl species (RCS) such as methylglyoxal (MGO) and glyoxal (GO) are common intermediates in protein damage, leading to the formation of advanced glycation end products (AGEs) through nonenzymatic glycation. (+)-Catechin, a natural plant extract from tea, has been evaluated for its ability in trapping GO and MGO. However, (+)-catechin is also reported to have both antioxidant ability and pro-oxidant properties. Until now, whether (+)-catechin can inhibit the formation of nonenzymatic glycation and the mechanism of the inhibition in nucleoprotein nonenzymatic glycation is still unclear. In the present study, histone H1 and MGO were used to establish an in vitro (100 mM phosphate buffer solution (PBS), pH 7.4, 37 °C) protein glycation model to study the trapping ability of (+)-catechin. Our data show that MGO caused dose-dependent protein damage, and the content of MGO-induced Schiff base formation was inhibited by (+)-catechin when the molecular ratio of catechin:MGO was 1:6. The formation of Nε-carboxymethyllysine (CML) was reduced significantly when the ratio of (+)-catechin and MGO was 1:1, which was similar to the inhibition effect of aminoguanidine (AG). The formation of CML under in vitro conditions can be inhibited by low concentration (12.5-100 µM) of (+)-catechin but not with high concentration (200-800 µM) of (+)-catechin. The reason is that the high concentration of (+)-catechin did not inhibit CML formations due to H2O2 produced by (+)-catechin. In the presence of catalase, catechin can inhibit MGO-induced CML formation. In conclusion, the trapping ability of (+)-catechin may be more effective at the early stage of nonenzymatic glycation. However, a high concentration (200-800 µM) of (+)-catechin may not inhibit the formation of CML because it induced the increase of H2O2 formation.

Role of autophagy and its signaling pathways in ischemia/reperfusion injury.

This study was conducted to investigate the mechanism of autophagy and its signaling pathways in ischemia/reperfusion injury (IRI). Pulmonary microvascular endothelial cells (PMVECs) were used to construct I/R models. The cells were then treated with autophagy inhibitor 3-MA and infected with adenovirus expressing Beclin 1-shRNA. The expression of CD31, LC3-II, Bcl-2, Bax, LC3-II, Beclin 1, AKT, p-AKT, AMPK and p-AMPK, apoptosis, cell viability and migration ability were determined. Over 95% isolated PMVECs were positive for CD31. The expression of LC3-II and Beclin 1 was up-regulated in I/R cells. 3-MA and Beclin 1 knockdown inhibited the expression of LC3-II and Beclin 1 and autophagosome formation. Autophagy induced by hypoxia was antagonistic against apoptosis, which increased after treatment with 3-MA and knockdown of Beclin 1. 3-MA and Beclin 1 knockdown downregulated and upregulated the expression of Bcl-2 and Bax, respectively. Apoptosis mediated by hypoxia and reperfusion-induced autophagy was reduced by 3-MA and Beclin-1 knockdown, which increased and reduced the expression of Bcl-2 and Bax, respectively, leading to significant decreased Bax/Bcl-2 ratio. In these cells, expression of p-AKT, p-AMPK and p-mTOR was up-regulated. After treatment with 3-MA and Beclin 1 knockdown, expression of p-AKT and p-AMPK was significantly reduced.

(+)-Catechin prevents methylglyoxal-induced mitochondrial dysfunction and apoptosis in EA.hy926 cells.

OBJECTIVE: To investigate whether (+)-catechin, a strong antioxidant, can prevent methylglyoxal (MGO)-induced cytotoxicity and its mechanism. METHODS: Cytotoxicity, apoptosis, reactive oxygen species (ROS) generation, hydrogen peroxide (H2O2) formation, mitochondrial membrane potential (MMP) and mitochondrial morphology were measured in EA.hy926 cells. RESULT: MGO (4 mM)-induced cytotoxicity was markedly inhibited by (+)-catechin (0.1-4 mM) in 24 h. 1 mM MGO-induced apoptotic cell death (44.7%) was significantly inhibited by 4 mM (+)-catechin (to 24.4%), 1 mM aminoguanidine (AG) (to 28.8%) or 4 mM N-acetylcysteine (NAC) (to 24.3%). (+)-Catechin (4 mM) or AG (4 mM) can inhibit the decrease of MMP induced by MGO (2-8 mM) in 3 h. (+)-Catechin (4 mM) or AG (4 mM) can inhibit MGO (4 mM)-induced mitochondrial swelling in 3 h. However, MGO (4 mM)-induced ROS and H2O2 generation was not prevented by (+)-catechin (4 mM). CONCLUSIONS: (+)-Catechin prevents MGO-induced cytotoxicity in EA.Hy926 cells through inhibiting apoptosis and mitochondrial damage.